Milk hygiene aspects of zoonos e s and non-zoonotic human pathogens

1. Milk hygiene aspects of zoonoses and non-zoonotic human pathogens

2. Microorganisms in the milk • Saprobes • 1, no illness (inhibit pathogens) • 2, no illness (organoleptic deviation – metabolism) • Pathogens • ILLNESS

3. Contamination of raw milk and milk products by pathogens • Primary/Intravital infection • Systemic diseases (salmonellosis, tuberculosis, brucellosis, listeriosis, Q fever) → excretion by milk • Mastitis • Subclinical: a few 104/ml • Clinical: 108/ml (pathogens! ↔ Somatic cell count)

4. Contamination of raw milk and milk products by pathogens • Secondary (fecal) contamination • Most common • During milking → hygiene deficiency of milking • Campylobacter, EHEC, L. monocytogenes • Other sources of contamination: air, dust, fixtures of stable, milking instruments, milk container, hand or clothes of stuff

5. Contamination of raw milk and milk products by pathogens • Recontamination (post-contamination) • After heat treatment • Sources of contamination: stuff or environment (water, air, dust) and fixtures, instruments • Especially dangerous (no way for killing)

6. Contamination of raw milk and milk products by pathogens • In general: • Pathogens can cause intravital infection in animals (mastitis) → no in herds or very rare? (tuberculosis, brucellosis; salmonellosis, listeriosis) • More common secondary contamination • WHO: 28 bacteria, viruses, unicellular pathogens can cause illness by milk consumption

7. Pathogens can contaminate raw milk • Bacteria • Gram +: Mycobacterium spp., S. aureus, L. monocytogenes, Bacillus cereus • Gram -: Brucella spp., Salmonella spp., Campylobacter spp., EHEC, Yersinia enterocolitica, Leptospira spp., Coxiella burnetti, Streptobacillus moniliformis • Viruses

8. Mycobacterium spp. • M. bovis, M. tuberculosis • Intravital/primary infection (without clinical symptoms of mastitis) • Food poisoning/ food born illness: Can live in fermented milk products made from raw milk (1-2 months)

9. Mycobacterium spp. • Food poisoning/ food-born illness: Can live in fermented milk products made from raw milk • Pasteurisation: to prevent infection byM. tuberculosis. (phosphatase enzymes inactived – pasteurisation – mycobacteria are killed)

10. Mycobacterium spp. • Prevention/treatment: pasteurisation and eradicationfrom herds • Milk: only from herds officially free from tuberculosis and brucellosis(Reg. 853/2004/EC) – protection from infected humans • Mycobacterium avium subsp. paratuberculosis (MAP)  paratuberculosis = Johne-disease (sheep, cow)  secondary contamination by faeces • (Chron-disease) • MAP can survive pasteurisation

11. Staphylococcus aureus • Most common contagious pathogens (mastitis – from animal to animal) • Primary/ intravital infection • Subclinical mastitis: 103–104/ml • Number of bacteria producing enterotoxin, can cause illness in humans: 106/ml • Milk have to be cooled ≤ 8°C (under 10°C – slow growing and no toxin production) • !Only 10-30% of the S. aureus strains can produce enterotoxin (human strains: 40-60%)!

12. Staphylococcus aureus • Source of contamination: clothes, hands of staff secondary contamination/ recontamination • Prevention/treatment:S. aureus can be killed by pasteurisation but the toxins are thermostable • Food poisoning/ food born illness: raw milk and fermented milk products made from raw milk

13. Listeria monocytogenes • Source of contamination: soil, gastrointestinal tract of animal secondary contamination • 1-100/ml milk • (in sheep, goat  mastitis – 104/ml) • Food poisoning/ food born illness: cheese (soft) made from raw milk (e.g. Roquefort, camembert)

14. Bacillus cereus • Source of contamination: soil • In raw milk max. 103-104/ml (Rarely cause illness) • Bacterial growth results in production of enterotoxin, and ingestion leads to two types of illness, diarrhoeal and emetic (vomiting) syndrome. • The emetic form: need 106-108/g bacteria for toxin production in the milk (at this Nr  remarkable spoilage of milk, circumstances not appropriate) • The diarrhoeal type: bacteria grow and produce toxin in the intestine but the vegetative form of bacteria are sensitive to acid in the stomach • Prevention/treatment: Appropriate cool storage

15. Brucella spp. • B. melitensis • Sheep, goat • Mediterranean area • In the EU more than 1000 case in the last few years • B. abortus (B. melitensis biovar. abortus) • Cow • Most of the European countries are officially free • Food poisoning/ food born illness: infected raw milk (sheep) and milk products made from raw milk (cheese, cottage cheese) • 100-100 000/ml

16. Brucella spp. • Resistance against heat or acid: low (60°C<, pH<4) • Pasteurised milk: ø • Fermented milk products: ø • Pasteurised cream: ø • Butter made from raw cream: virulent for 30 days • Soft and semi-hard cheese made from raw milk: + • MILK: no organoleptic changes, somatic cell count (SCC) ↑, Langhans-type giant cells • Milk: only from herds officially free from tuberculosis and brucellosis (Reg. 853/2004/EC)

17. Salmonella spp. • Role of milk in Salmonella-induced food born illness is low • Rarely cause septicaemia and (as a consequence) mastitis (intravital infection) • Milk: SCC ↑, Cl-↑, flakes, greyish-brown, putrid • Secondary contamination (most common) during milking, handling • Food poisoning/ food born illness: • Pasteurised milk: ø • Fermented milk products: ø • Butter, soft and semi-hard cheese made from raw milk: viable for 30 days

18. Campylobacter spp. • C. jejuni, C. coli • (intravital infection) • Secondary contamination by faeces • Source of contamination: environment, milking equipments, hand or clothes of stuff • Thermophyl  Summer (longer storage without cooling)  food borne infection • Prevention/treatment: cooling (inhibit growing) and pasteurisation

19. Enterohaemorrhagic E. coli (EHEC)

20. Enterohaemorrhagic E. coli (EHEC) • (Intravital infection – mastitis 0-2%) • Secondary contamination by faeces • Higher tolerance to acid  fermented milk products (virulent) • Infectious dose is low (10-100) • Food poisoning/ food born illness: raw milk, products made from raw milk (Mould-ripened soft cheeses) • Prevention/treatment: pasteurisation

21. Yersinia enterocolitica • Secondary contamination by faeces • During milking, handling • Prevention/treatment: pasteurisation

22. Leptospira spp. • Bovine leptospirosis - worldwide incidence • Intravital infection  mastitis • Milk: yellow, dense consistency • At 4°Craw milk: viable for 2 month • Prevention/treatment: pasteurisation, low pH

23. Coxiella burnettii • Worldwide common, resistant bacterium • Primary/intravital infection (no symptoms) • At 4°C raw milk: viable for months • Sensitive to low pH • Can survive flash and traditional pasteurisation at lower range (HTST – 72°C, 15sec; LTLT – 62°C, 30min)

24. Rat-bite fever: Streptobacillus moniliformis • Rats are the main reservoir of the organism • Human infection is usually acquired either from a rat bite/scratch, handling infected rats, or, • In the case of the form of disease known as Haverhill fever, ingestion of milk or water contaminated with the organism (via rat urine)

25. Viruses • 1. Source of contamination: human faeces, (poor personal hygiene)  secondary contamination • Prevention/treatment: pasteurisation • 2. intravital/primary infection

26. Viruses • Tick-borne encephalitis • Food poisoning/ food born illness: raw milk, products made from raw milk (soft cheese) • Rabies • Can be detected in milk but no evidence of human infection • Foot and mouth disease • Before symptoms (asymptomatic) milk can be infected • Food poisoning/ food born illness: raw milk, and cream

27. Chemical contamination of milk

28. Chemical contaminants • Intravital contamination • Treatment – pharmaceuticals • Environment • Secondary contamination • Environment • Milking, milkprocessing (e.g. additives)

29. Legislation • Regulation 1881/2006/EC (mycotoxins, heavy metals, dioxin-like materials, Polycyclic Aromatic Hydrocarbons (PAH) • Regulation 2377/90/EEC (residue limits of pharmacologically active substances) • Regulation 2218/89/EEC (radioactive contamination of foodstuffs)

30. Most common substances contaminating milk • Pharmaceuticals • Pesticides • Contaminants of environmental origin (heavy metals, polychlorinated organic materials) • Mycotoxins • Biogenic amines • Radioactive substances

31. Milk hygienic importance of pharmacology residues • Treatment of mastitis • Intramammal (direct) application • Through the milk-blood barrier from the blood (lypophyl, non-ionic, less-binding to blood proteins)

32. Milk hygienic importance of pharmacology residues • Excretion by milk • Some products/pharmaceuticals are not permitted to use for treatment of lactating cattle (e.g.: avermectins) • For other products withdrawal time have to be determined (based on MRL value) • MRL values of milk lower than of other edible tissues

33. MRL values of milk and other edible tissues

34. Milk hygienic importance of veterinary drug residues • Unwholesome effects: Allergic reaction • 10 IU penicillin derivate (1 IU benzylpenicillin = 0.6 μg) • In the case of antibiotics, residues can inhibit starter cultures in the production of yoghurt, cheese and other fermented milk products • 0.01 IU/ml inhibit lactobacillus in fermentation

35. Detection of drug residues • Screening test: • inhibition method • Sample + bacterium suspension (Bacillusstearothermophylus var. calidolactis) + indicator  nutrient media • Growing: - • Signs of metabolism: - • Confirmative test: • ELISA-methods, • Chromatographic, • Spectroscopic, • Electrochemical analysis • Milk containing residue is NOT ACCEPTABLE for human consumption and for further processing and as feed for animals

36. Residues of pesticides • Chemical pest control of plants • Pest and insect control •  Contaminated feed and drinking water • Excretion into milk • Typical characteristics/properties of pesticidesoccurring in milk: • Lipophyl • Easy absorption from intestinal tract • Poorly metabolised • Passive diffusion through the blood-milk barrier to milk • E.g.: chlorinated hydrocarbons, like DDT-group (DDT-dichlorodiphenyltrichloroethane, dieldrin, aldrin, etc.)

37. Residues of pesticides • In most countries of Europe were banned 40 years ago • Very persistent (Persistent Organic Pollutants – POPs) but now, the concentration in the environment is low  in concentrations above residue limit can not be detected • Lipophylic  in cream or butter the concentration can be higher • Accumulate in mother’s milk  higher (30 times) concentration and can be detected also today!

38. Other important pesticides • Organic phosphoric acid esters • Pyrethrins, pyrethroids • Fast metabolisation  not present in milk in considerable concentration.

39. The maximum residue limits (MRL) for milk and milk products • * banned active substance

40. Contaminants of environmental origin • Toxic heavy metals • Cadmium, lead, methyl-mercury • Polychlorinated organic pollutants • Dioxins, furans, biphenyls

41. Contaminants of environmental origin - Toxic heavy metals • Cadmium (Cd), and lead (Pb) • Poor absorption from intestinal tract (<10%) • Accumulation in the organism • Can be present in the milk • Not considerable (EU: 3-5% of Cd taken by humans is from milk or milk products) • Methyl-mercury • Good absorption from intestinal tract • Good passive transport through the blood-milk barrier  Can be present in the milk (at low conc.) • 10% of Hg taken by humans is from milk

42. Maximum metal content of food of animal origin • x: no criteria

43. Contaminants of environmental origin - Polychlorinated organic pollutants • Dioxins, furans, biphenyls • Lipophyl • Persistent • Are disposed to accumulate in food chain • Can be present in food (among others milk, butter, cream) • Generally lower than the criteria (Reg. 1881/2006/EC)

44. Mycotoxins • Contaminated feed  animal  milk human • Aflatoxins • Ochratoxin • Zearalenone

45. Mycotoxins • Aflatoxin B1 and B2 in lactating cow by oxidation is transformed to 4-hydroxy-metabolites named „milk toxin” = aflatoxin M1 and M2 • 1-3% of the toxin of feed is excreted by milk • The aflatoxin M1 can not be inactivated by pasteurisation and in fermented milk products is quite stable • Genotoxic carcinogen effect of aflatoxin M1 is a little bit lower than that of B1

46. Mycotoxins • Ochratoxin A (OTA) can be present potentially in the milk, but mainly in less toxic metabolite form (OTAα) – no criteria • Zearalenone (ZEN) can be present potentially in the milk, but mainly in metabolite form (α and ß-ZEN) – no health risk for humans

47. Biogenic amines • Biogenic amines are formedfrom amino acids by microbial decarboxylation • Low concentration in milk but higher in some cheese

48. Biogenic amines in cheeses

49. Biogenic amines • Bacteria can produce biogenic amines (decarboxylase enzyme).They can be found in raw milk in higher amount→ The amount of biogenic amines is higher in cheese produced from raw milk. • Biogenic amines produced by bacteria can contaminated milk during cheese production (from instruments and equipment) • Bacteria of the starter cultures also can produce biogenic amines.

50. Biogenic amines • The requirednumber of bacteria is 106 cell/g cheese • Enterobacteria can not proliferate to reach this number in case of hygienic production technology →Lactobacillus and Enterococcus species are the main biogenic amine producers.