Dairy microbiology

In short

Dairy microbiology is the study of bacteria, yeasts, moulds, bacteriophages and their enzymes in milk and milk products — those that spoil milk, those that cause disease, and those deliberately added as starters and probiotics. It decides shelf life, safety, heat-treatment design and the price a dairy pays a farmer, because raw-milk grading in India is largely a microbial judgement (MBRT, SPC, coliform count). It is a core paper in every B.Tech Dairy Technology curriculum (dairy microbiology, dairy bacteriology lab, starter cultures) and is examined in ICAR/ASRB JRF-SRF, GATE XL/Food Technology and university practicals. In placement rounds at Amul (GCMMF member unions), Mother Dairy, Nestlé, Hatsun, Heritage, Dodla, Schreiber Dynamix and Britannia, QA and production interviews open with MBRT grading, phosphatase failure, post-pasteurisation contamination and starter troubleshooting.

Explained

Microflora of raw milk — sources and groups

• Milk inside a healthy udder is nearly sterile; the teat canal adds a few hundred micrococci and streptococci. Typical aseptically drawn milk: 10²–10³ CFU/mL.
• Sources of contamination: udder exterior and dung, bedding and air, unclean cans and milking machine (biofilms in rubber parts and dead ends are the worst single source), water, flies, the milker's hands, and the transport chain.
• Groups by growth temperature: psychrotrophs (grow below 7 °C; optimum 20–30 °C), mesophiles (20–45 °C), thermophiles (optimum 50–55 °C), thermoduric (survive pasteurisation), thermophilic spore formers (survive and grow in UHT pre-heat and evaporator sections).
• Psychrotrophs — *Pseudomonas fluorescens*, *Ps. fragi*, *Acinetobacter*, *Flavobacterium*, *Aeromonas*. They dominate refrigerated raw milk (>75% of the flora after 48–72 h at 4 °C) and secrete heat-stable lipases and proteases that survive UHT; these cause rancid and bitter flavours and age gelation in UHT milk months later. Hence "cold storage does not undo bad hygiene".
• Lactic acid bacteria (LAB) — *Lactococcus lactis* subsp. *lactis* and *cremoris*, *Streptococcus thermophilus*, *Lactobacillus delbrueckii* subsp. *bulgaricus*, *Lb. helveticus*, *Leuconostoc mesenteroides* subsp. *cremoris*, *Lacticaseibacillus casei* (several *Lactobacillus* species were reclassified into new genera in 2020 — older textbooks use the old names).
• Coliforms — *Escherichia coli*, *Klebsiella*, *Citrobacter*, *Enterobacter*: Gram-negative, lactose-fermenting with gas, killed by pasteurisation. Therefore they are indicators of faecal/unhygienic contamination in raw milk and of post-pasteurisation contamination in processed milk, not a pasteurisation-adequacy test by themselves.
• Spore formers — *Bacillus cereus* (sweet curdling and bitty cream in pasteurised milk; also a toxin producer), *B. licheniformis*, *B. subtilis*, *Geobacillus stearothermophilus* and *Anoxybacillus* (flat-sour in canned/UHT products), *Clostridium tyrobutyricum* (late blowing of Gouda/Swiss-type cheese from silage), *C. sporogenes*, *C. botulinum* (risk in low-acid canned dairy).
• Yeasts and moulds — *Kluyveromyces marxianus*, *Candida*, *Geotrichum candidum*, *Penicillium*, *Aspergillus*, *Mucor*, *Rhizopus*: spoil paneer, khoa, butter, cheese and sweetened condensed milk; they tolerate acid, salt and low water activity, so they spoil products bacteria cannot.

Spoilage and defects — which organism causes what

• Souring/acid coagulation: LAB and coliforms fermenting lactose to lactic acid; milk clots when pH falls to about 4.6 (casein isoelectric point).
• Sweet curdling: *B. cereus* rennet-like protease clots milk without acid.
• Proteolysis / bitterness / putrefaction: *Pseudomonas*, *Proteus*, *Bacillus*; bitter peptides from casein breakdown.
• Lipolysis / rancidity: microbial and native lipases release butyric and caproic acids; aggravated by agitation, foaming and repeated pumping of raw milk.
• Ropiness: *Alcaligenes viscolactis*, *Klebsiella*, exopolysaccharide-producing LAB (desirable in dahi/yoghurt body, a defect in market milk).
• Gas and blowing: coliforms, yeasts, clostridia (late blowing in hard cheese).
• Colour defects: blue (*Ps. syncyanea*), yellow (*Ps. synxantha*), red (*Serratia marcescens*, *Brevibacterium erythrogenes*).
• Product-specific: ghee — oxidative, not microbial (low moisture); butter — surface taint by *Ps. putrefaciens*, moulds; paneer and khoa — staphylococci, coliforms, yeasts and moulds, shelf life 1–3 days at ambient and about a week at 5–7 °C; sweetened condensed milk — osmophilic yeasts and *Micrococcus*; milk powder — no growth at a_w ≈ 0.2 but survivors of *Salmonella*, *Cronobacter sakazakii* and *B. cereus* persist for months (a critical infant-formula hazard).

Milk-borne pathogens and public health

• Zoonotic (from the animal): *Mycobacterium bovis* and *M. tuberculosis* (pasteurisation's historical target), *Brucella abortus* and *B. melitensis* (brucellosis — also an occupational risk for dairy workers), *Coxiella burnetii* (Q fever; the most heat-resistant non-spore pathogen, which sets the modern HTST benchmark), *Streptococcus agalactiae*, *Staphylococcus aureus* (mastitis).
• From handling/environment: *Salmonella* spp., enterohaemorrhagic *E. coli* O157:H7, *Listeria monocytogenes* (grows at 0–4 °C, high-risk in soft cheese and ice cream), *Campylobacter jejuni*, *Shigella*, *Yersinia enterocolitica*, *Cronobacter sakazakii* (infant formula), *B. cereus*.
• Toxins: staphylococcal enterotoxin is heat-stable and survives pasteurisation — so toxin formed in abused raw milk stays a hazard after heating. *B. cereus* cereulide likewise.
• Aflatoxin M1 — the hydroxylated metabolite excreted in milk when the animal eats aflatoxin B1-contaminated feed (groundnut cake, maize); not destroyed by pasteurisation or boiling. FSSAI prescribes a limit for aflatoxin M1 in milk (commonly cited as 0.5 µg/kg; verify the current figure in the Food Safety and Standards (Contaminants, Toxins and Residues) Regulations, 2011 as amended, since limits are revised).
• Antibiotic residues: after mastitis therapy, residues both endanger consumers and inhibit starter cultures (slow/dead vat). Observe the labelled withdrawal/withholding period; test by delvotest/charm or disc-assay. Residue limits are specified in the FSSAI contaminants regulations.
• Somatic cell count (SCC): leucocytes shed in mastitis; high SCC means lower casein and lactose, poorer cheese yield and keeping quality. Bulk-milk SCC above roughly 2–4 lakh cells/mL signals subclinical mastitis in the herd; the California Mastitis Test (CMT) is the field screen.

Testing, grading and the Indian regulatory frame

1. Sampling: mix thoroughly, sterile dipper/plunger, sterile bottle, chill to ≤4 °C, test within hours; for a composite, sample proportional to volume.
2. Standard Plate Count (SPC): pour plate in Plate Count Agar (standard methods agar), 32 °C/48 h (or 30 °C/72 h); report CFU/mL as count × dilution factor. Countable range 25–250 (ISO/IDF uses 15–300).
3. Direct Microscopic Count (DMC): 0.01 mL smeared over 1 cm², methylene blue/Newman-Lampert stain; counts living + dead cells and clumps, hence higher than SPC; also shows leucocytes and cell morphology. Microscopic factor = 1/(area of field × 0.01).
4. MBRT (methylene blue reduction test): 10 mL milk + 1 mL standard methylene blue, incubate at 37 °C, note time to decolourise. Bacterial dehydrogenases lower redox potential. Longer time = better milk. Cheap, no incubator-dependent plating; limitation — insensitive to psychrotrophs and thermodurics, and leucocytes/dissolved oxygen interfere.
5. Resazurin test: same principle, colour goes blue → purple → pink → white; read at 10 min (1-hour or 10-minute test) and match a disc comparator — faster than MBRT, used for quick payment decisions at chilling centres.
6. Clot-on-boiling (COB) and alcohol (68–75%) test: platform rejection tests for high-acid or unstable milk; titratable acidity (% lactic acid) supports them.
7. Coliform count: violet red bile agar (VRBA) pour plate with overlay, 32–37 °C/24 h, dark red colonies ≥0.5 mm with bile precipitate; confirm gas in BGLB. MPN method for low counts. *E. coli* confirmed on EMB/ EC broth at 44.5 °C or by IMViC.
8. Phosphatase test: alkaline phosphatase in milk is destroyed just beyond HTST conditions, so a negative phosphatase = adequate pasteurisation + no raw-milk admixture. Fluorometric (Fluorophos) and Scharer tests are used; a false positive can come from reactivation in cream/UHT products, so a positive result is confirmed.
9. Turbidity test: confirms sterilisation/adequate heating of sterilised milk — a negative turbidity test means whey proteins were fully denatured.
10. Other: peroxidase (Storch) test shows heating above about 80 °C; thermoduric count = count after laboratory pasteurisation at 63 °C/30 min; spore count = after 80 °C/10 min heat shock; preliminary incubation (PI) count at 12.8 °C/18 h targets psychrotrophs; ATP bioluminescence swabs and Petrifilm/impedance/flow cytometry (Bactoscan) for rapid plant QC; PCR/ELISA for pathogen confirmation.
11. Regulatory frame: the Food Safety and Standards Act, 2006 with the FSS (Food Products Standards and Food Additives) Regulations, 2011 (compositional standards) and the FSS (Food Safety and Standards relating to Microbiological Standards) Regulations — microbiological limits for milk and milk products were consolidated in 2020 and are periodically amended, so quote current limits from the FSSAI site rather than from memory. Schedule 4 of the FSS (Licensing and Registration) Regulations prescribes hygiene and GMP/GHP for dairy units, and FSSAI's FoSTaC training and RUCO/ Hygiene Rating schemes apply to processors. BIS standards exist for pasteurised milk, milk powders and methods of test (IS 1479 series, methods of sampling and test for dairy industry); several older IS numbers have been revised or withdrawn, so verify the current edition. Export of dairy products is cleared through APEDA/EIC with plant approval.

Heat treatment, thermal death kinetics and plant hygiene

• Thermisation 57–68 °C/15 s — reduces psychrotrophs before storage. LTLT (holder/batch) 63 °C/30 min. HTST 72 °C/15 s. Higher-heat shorter-time up to 89–100 °C for ESL. UHT 135–150 °C/2–4 s plus aseptic packing (commercially sterile, ambient shelf life). In-bottle sterilisation 115 °C/15 min. Boiling of milk at the household level is common in India and far harsher than HTST.
• Design basis: HTST gives at least a 5-log kill of *C. burnetii* and *M. tuberculosis* but is not sporicidal and does not inactivate heat-stable microbial lipases/proteases or Staph enterotoxin.
• D value: time at a constant temperature for a 1-log (90%) reduction. z value: temperature rise needed to cut D ten-fold (about 4–5 °C for vegetative cells, 7–10 °C for spores). F value: equivalent heating time at a reference temperature (F₀ at 121.1 °C, z = 10 °C for clostridial spores).
• Post-pasteurisation contamination (PPC) is the number one cause of short shelf life in Indian pouch milk: cracked plates (detect by raw-milk leakage/phosphatase), filler nozzles, poor pouch film, condensate. Diagnose with coliform count, PI count and plant swabs — a pasteurised sample with coliforms means PPC, not under-pasteurisation.
• CIP: rinse → hot alkali (1–2% NaOH, 70–80 °C) to remove protein/fat → rinse → acid (0.5–1% HNO₃/phosphoric) to remove milkstone → rinse → sanitise (hypochlorite 100–200 ppm, peracetic acid, hot water 85 °C, or steam). Rotate sanitisers; validate with swab/rinse counts.
• Biofilms in gaskets, pump seals and dead legs re-seed the line continuously; they resist sanitisers hundreds of times better than free cells.
• HACCP/ISO 22000: pasteurisation is the classic CCP with critical limits of temperature and holding time, continuous recording thermograph and flow diversion valve (FDV) as the control; monitoring is the chart plus phosphatase; corrective action is re-pasteurisation.

Starter cultures, fermentation and probiotics

• Definition: a starter is a selected culture of LAB (sometimes with yeasts/moulds) added to milk to produce acid, flavour, texture and safety by competition.
• Classification: mesophilic (optimum 20–30 °C) — *Lc. lactis* subsp. *lactis*/*cremoris* for dahi, cultured butter, cheddar, cottage cheese; thermophilic (37–45 °C) — *S. thermophilus* + *Lb. bulgaricus* for yoghurt, mozzarella, Swiss/Emmental (with *Lb. helveticus*, *Propionibacterium freudenreichii* for eyes). Also single-strain, multiple-strain, mixed-strain and DVS (direct vat set) frozen/freeze-dried concentrates, which have replaced bulk mother-cultures in most Indian plants.
• Functional types: homofermentative (>85% lactic acid from lactose — *Lactococcus*, *S. thermophilus*, *Lb. bulgaricus*) vs heterofermentative (lactic acid + CO₂ + acetate/ethanol — *Leuconostoc*, *Lb. fermentum*); citrate-fermenting (aroma/"D"/"LD") cultures producing diacetyl, the buttery aroma of dahi, buttermilk and butter.
• Yoghurt symbiosis (protocooperation): *S. thermophilus* grows first, its proteinase-free growth is helped by peptides and amino acids (valine, glycine, histidine) released by *Lb. bulgaricus* proteinases; *S. thermophilus* in turn produces formate, pyruvate and CO₂ that stimulate the lactobacillus. Ratio about 1:1; incubate 42–43 °C to pH 4.5.
• Indian fermented products: dahi (mesophilic or mixed, set at 25–30 °C or 37–42 °C to 0.7–0.9% acidity), misti dahi/mishti doi (caramelised, sweetened, *S. thermophilus*-led), lassi and chhash, shrikhand (chakka from dahi), yoghurt (set, stirred, drinking, Greek), cheeses (*Lactococcus* + rennet or microbial coagulant; *Penicillium roqueforti*/*camemberti* for mould-ripened), cultured butter, kefir and koumiss (LAB + lactose-fermenting yeast, ethanol and CO₂), and traditional products such as chhurpi and dahi-based kadhi.
• Starter failure — causes: bacteriophage attack (the classic one), antibiotic/sanitiser residues, high free fatty acids, agglutinins in raw milk, wrong incubation temperature, mastitic or high-SCC milk, natural inhibitors (lactoferrin, lactoperoxidase system), too little inoculum, repeated subculturing causing strain drift. Phage control: rotate unrelated strains, phage-resistant/phage-inhibitory media, aseptic bulk-starter tanks, DVS use, chlorine fogging and strict separation of whey and starter areas.
• Probiotics: live microbes that confer a health benefit at adequate intake — *Lacticaseibacillus rhamnosus* GG, *Lactobacillus acidophilus*, *Bifidobacterium* spp., *Lactiplantibacillus plantarum*, *Saccharomyces boulardii*. Indian examples: Yakult (*Lc. casei* Shirota), Amul and Mother Dairy probiotic dahi/lassi, NDRI/NDDB strains. FSSAI's Food Safety and Standards (Health Supplements, Nutraceuticals, Food for Special Dietary Use…) Regulations, 2016 list permitted strains and generally require a viable count of the order of 10⁸ CFU per serving through shelf life — check the current schedule and amendments. Prebiotics (inulin, FOS, GOS, lactulose) feed them; synbiotic = both. Bacteriocins: nisin from *Lc. lactis*, pediocin — natural preservatives (nisin is a permitted additive in specified products).

Key points

• Udder milk - nearly sterile (10²–10³ CFU/mL); almost all contamination is from equipment, handling and water, not the cow.
• Psychrotrophs - grow at ≤7 °C and leave heat-stable lipases/proteases that survive UHT; they cause rancidity, bitterness and UHT age gelation.
• Thermoduric vs thermophilic - thermoduric survive pasteurisation; thermophilic prefer 50–55 °C and multiply in regenerator/evaporator hot spots.
• Coliforms - indicator of unhygienic or faecal contamination in raw milk and of post-pasteurisation contamination in heated milk; killed by HTST.
• Phosphatase test - negative means pasteurisation was adequate and no raw milk was mixed in; turbidity test confirms sterilisation.
• MBRT/resazurin - dye-reduction (redox) tests, quick grading at collection centres; longer decolourisation time = better milk; weak on psychrotrophs.
• SPC - pour plate, 32 °C/48 h, countable 25–250 colonies; CFU/mL = colonies × dilution factor.
• DMC > SPC because the microscope counts dead cells and clumps too.
• HTST 72 °C/15 s - benchmark set by *Coxiella burnetii*; not sporicidal and does not destroy staphylococcal enterotoxin.
• D value - time for 90% (1-log) kill; z value - °C rise that reduces D ten-fold; F₀ - equivalent minutes at 121.1 °C.
• B. cereus - sweet curdling and bitty cream in pasteurised milk; spores survive pasteurisation and germinate in cold storage.
• Clostridium tyrobutyricum - late blowing of hard cheese from silage-fed milk; controlled by bactofugation, nitrate or lysozyme.
• Listeria monocytogenes - grows at refrigeration temperature; the key hazard of soft cheese, ice cream and RTE dairy.
• Aflatoxin M1 and antibiotic residues - heat-stable, carried from feed or treatment; residues also kill starters.
• Starter culture - selected LAB for acid, flavour and texture; mesophilic (20–30 °C) vs thermophilic (37–45 °C); DVS has displaced bulk mother cultures.
• Yoghurt protocooperation - *S. thermophilus* + *Lb. bulgaricus* feed each other (peptides ↔ formate/CO₂) at 42–43 °C.
• Bacteriophage - the commonest cause of slow vats; controlled by strain rotation, phage-inhibitory media and hygiene.
• Milk clots at pH ≈ 4.6 - the isoelectric point of casein; dahi is set to about 0.7–0.9% lactic acid.
• Legal frame - FSS Act, 2006; FSSAI microbiological standards (consolidated 2020, amended since); Schedule 4 hygiene; pasteurisation is the textbook HACCP CCP.

Formulas and examples

Example 1 — SPC from plate counts. 1 mL of a 10⁻⁴ dilution gives 68 colonies.

• SPC = 68 × 10⁴ ÷ 1 = 6.8 × 10⁵ CFU/mL.
• If the 10⁻³ plate gave 540 colonies, it is above the 250 limit and is discarded; only the in-range plate is reported.

Example 2 — weighted average over two dilutions. 1 mL plated; 10⁻³ plates give 232 and 244; 10⁻⁴ plates give 33 and 28.

• ΣC = 232 + 244 + 33 + 28 = 537; n₁ = 2, n₂ = 2, V = 1 mL, d = 10⁻³.
• N = 537 / [1 × (2 + 0.2) × 10⁻³] = 537 / (2.2 × 10⁻³) = 2.44 × 10⁵ → report 2.4 × 10⁵ CFU/mL.

Example 3 — generation time in warm milk. Milk held at 30 °C rises from 5 × 10⁴ to 2 × 10⁷ CFU/mL in 5 hours.

• log₁₀(2 × 10⁷) = 7.301; log₁₀(5 × 10⁴) = 4.699; difference = 2.602.
• n = 3.322 × 2.602 = 8.64 generations; g = 300 min / 8.64 = 34.7 min.
• Take-away: an unchilled 5-hour delay in collection multiplied the count 400-fold — chilling to 4 °C within 2–3 h is the single most effective control.

Example 4 — D and z values, log reduction. *C. burnetii* has D₆₃ = 0.6 min with z = 4.5 °C.

• At 72 °C: D₇₂ = 0.6 × 10^[(63 − 72)/4.5] = 0.6 × 10⁻² = 0.006 min = 0.36 s.
• HTST holding 15 s → log reduction = 15 / 0.36 = 41.7 logs — an enormous margin for this organism.
• Spores of *G. stearothermophilus* with D₁₂₁ = 4 min: a 12-log kill needs 48 min at 121 °C, which is why pasteurisation cannot be sporicidal.

Example 5 — titratable acidity of dahi. 10 g dahi needs 9.5 mL of 0.1 N NaOH to the phenolphthalein end point.

• Acidity = (9.5 × 0.009 × 100)/10 = 0.855% lactic acid — within the usual 0.7–0.9% target for good dahi; further holding at 30 °C would over-acidify and cause whey separation.

Where it is used

• Milk chilling centres and BMCUs of cooperatives (Amul member unions, Mother Dairy, Nandini/KMF, Milma, Aavin, Verka, Saras): MBRT/resazurin and COB at reception decide acceptance and grade-based payment to farmers.
• Liquid-milk plants: pasteuriser CCP monitoring, phosphatase, coliform and PI counts, PPC hunting, CIP and swab validation — the daily work of a QA/QC officer (typical entry pay roughly ₹3.5–6 lakh per annum for a B.Tech Dairy graduate, higher in MNCs).
• Fermented-product and cheese plants: starter propagation or DVS handling, phage control, strain rotation and sensory/acidity monitoring — Amul, Britannia, Nestlé, Schreiber Dynamix, Go, Epigamia, ITC; probiotic dahi and lassi lines.
• Powder, infant-formula and UHT plants: thermophilic spore control in evaporators, *Cronobacter* and *Salmonella* environmental monitoring programmes, aseptic-line sterility testing.
• Regulators and labs: FSSAI-notified/NABL-accredited food labs, state food safety officers, EIA/EIC for dairy exports, BIS testing; R&D and teaching at NDRI Karnal, NDDB, SMC College Anand, and the dairy science colleges under state agricultural universities.
• Animal-side roles: mastitis and SCC control, clean-milk-production training of farmers, feed-mould and aflatoxin management in milk unions' extension wings.

Interview questions

• Q: Is milk drawn from a healthy udder sterile? A: Practically but not absolutely — it carries about 10²–10³ CFU/mL of micrococci and streptococci from the teat canal; the heavy load comes from equipment, hands, water and air.
• Q: A pasteurised milk sample is phosphatase-negative but has 50 coliforms/mL. What happened? A: Pasteurisation was adequate, so this is post-pasteurisation contamination — suspect the filler, pouch film, cooler section after the holding tube, or condensate/biofilm downstream.
• Q: Why doesn't chilling solve a hygiene problem? A: Cold selects psychrotrophic *Pseudomonas*, which secrete heat-stable lipases and proteases that survive even UHT and later cause rancidity, bitterness and age gelation.
• Q: Which organism sets the HTST time–temperature combination? A: *Coxiella burnetii*, the most heat-resistant non-sporing milk pathogen; the earlier benchmark was *Mycobacterium tuberculosis*.
• Q: Why is DMC higher than SPC for the same milk? A: DMC counts dead as well as live cells and each clump's individual cells, whereas SPC counts only viable units capable of forming a colony, and a clump gives one colony.
• Q: What is the principle of MBRT and what does a 30-minute decolourisation mean? A: Bacterial dehydrogenases lower the redox potential and reduce methylene blue to its colourless form; decolourisation within about 30 minutes indicates a very high bacterial load — poor-grade milk, usually rejected or diverted.
• Q: Pasteurised milk curdles sweetly without souring. Diagnose. A: *Bacillus cereus* — its rennet-like protease clots casein without acid; the spores survived pasteurisation and germinated during cold storage.
• Q: Your dahi vat is slow and the acidity has stalled. List causes in order of likelihood. A: Bacteriophage attack, antibiotic or sanitiser residue in the milk, wrong incubation temperature, insufficient or weakened inoculum, mastitic/high-SCC milk, natural inhibitors.
• Q: How do *S. thermophilus* and *Lb. bulgaricus* help each other? A: The lactobacillus proteinases release peptides and amino acids for the streptococcus, which returns formate, pyruvate and CO₂ that stimulate the lactobacillus — protocooperation, run at 42–43 °C.
• Q: Does boiling make aflatoxin-contaminated milk safe? A: No — aflatoxin M1 is heat-stable; control is at the feed end, by preventing aflatoxin B1 in cattle feed and testing milk against the FSSAI limit.
• Q: Difference between thermoduric and thermophilic bacteria? A: Thermoduric organisms merely survive pasteurisation (measured by count after 63 °C/30 min); thermophilic organisms actively grow at 50–55 °C, for example in regenerator and evaporator sections.
• Q: Why do hard cheeses blow late, and how is it prevented? A: *Clostridium tyrobutyricum* spores from silage ferment lactate to butyric acid and CO₂ after weeks of ripening; prevented by silage-free feeding, good milking hygiene, bactofugation or microfiltration, permitted nitrate or lysozyme.
• Q: Which single lab result best predicts the shelf life of pouch milk? A: The preliminary-incubation or psychrotrophic count on the finished pack, backed by coliform count — both reflect psychrotrophs and PPC, which govern spoilage, rather than total load alone.

Common mistakes

• Reporting an SPC from a plate with fewer than 25 or more than 250 colonies instead of choosing the in-range dilution, or forgetting to multiply by the dilution factor.
• Treating coliform absence as proof of safety, or a coliform-positive pasteurised sample as under-pasteurisation when it is almost always post-pasteurisation contamination.
• Saying pasteurisation "sterilises" milk — it is neither sporicidal nor able to destroy heat-stable enzymes and staphylococcal enterotoxin.
• Confusing D and z values, or using temperature in °F/Kelvin inconsistently in the D_T = D_ref × 10^[(T_ref − T)/z] relation.
• Mixing up thermoduric, thermophilic and psychrotrophic groups, and assuming refrigeration stops all growth (it does not stop *Listeria* or *Pseudomonas*).
• Calling MBRT a measure of psychrotrophs or of pasteurisation efficiency — it is a crude dye-reduction load test on raw milk only.
• In practicals: inverting pour plates before the agar sets, pouring agar above 48 °C and killing cells, ignoring the VRBA overlay, or incubating plates at the wrong temperature.
• Quoting withdrawn IS numbers or outdated FSSAI microbiological limits from old notes instead of the current consolidated standards.

Related topics

Dairy quality and FSSAI standards · Market milk processing and pasteurisation · Fermented milk products and cheese technology · Dairy plant sanitation, CIP and HACCP · Food microbiology and fermentation technology · Milk chemistry and composition