E CONTROL (reduced-fat spread)

Summary Signiﬁcant hazards a Although there are no reported cases of food-borne illness associated with

consumption of reduced-fat spreads, signiﬁcant hazards to which adequate control measures need to be established are:

r Salmonella spp.

E. coli O157:H7. r L. monocytogenes.

Control measures

Initial level (H 0 ) r Use appropriate ingredients (source from approved supplier and/or

pasteurize ingredients in-house before use). r Use potable water from a reliable source.

Reduction (ΣR)

r Use preservatives where appropriate. r Pasteurise water or pre-emulsion phase; vulnerable formulations may

require in-line pasteurization of the complete emulsion. Increase (ΣI)

r Use appropriate product structure: i.e. an appropriately ﬁne water-in-oil dispersion to prevent or limit growth of microorganisms.

r Avoid (re-)contamination; very vulnerable formulations may require

speciﬁc facilities for aseptic handling. r Use suitable, hygienic equipment and process hygienically (incl. proper

cleaning). r Keep end-product refrigerated (closed and open shelf-life); store dry and

prevent condensation.

Testing

r Ingredient specification and water quality should be verified. r Interim and finished product should be tested regularly to verify process

performance. r Routine end product testing is not advised when a stable product

formulation/structure is used and process control is adequate.

Spoilage

r Generally, spoilage problems are controlled by using a stable formulation and an appropriate product composition, i.e. correct pH, clean

ingredients, and appropriate water dispersion. Special attention needs to

be given to the quality (speciﬁcations) of the stabilizing agents. a In particular circumstances, other hazards may need to be considered.

Control measures. Considering the relative vulnerable nature of a reduced-fat spread formulation, a process design based on a cold manufacturing process alone may not be sufﬁcient to achieve adequate microbiological product safety and stability. The use of suitable (pasteurized) raw materials is essential, in-line pasteurization is commonly employed and recontamination during processing (incl. packaging) should be avoided. In the case of very sensitive products, refrigerated distribution should be considered.

Raw materials. To meet the right microbiological standards, raw materials must be selected as in the case of margarine. Special attention must be given to materials that are used to stabilize the aqueous

MICROORGANISMS IN FOODS 6

contain only low numbers of thermophilic spore-formers because of the way it is manufactured, but these can grow rapidly, for instance in a stock solution of gelatin in water at 50–55 ◦

C. A low amount of potassium sorbate may help preserve the gelatine stock solution. Starches may contain Salmonella spp. and occasionally contain high numbers of bacterial spore-formers, including Bacillus (particularly B. cereus in, for instance, rice starch). Typical microbiological speciﬁcations for starches and

gums are aerobic plate count <10 4 cfu/g, Enterobacteriaceae <100 cfu/g, yeasts and molds <500 cfu/g,

B. cereus <1000 cfu/g, absence of infectious pathogens. Because the ﬁnished product may include bacterial spores, its composition and the physical structure should not allow their outgrowth.

Processing. Although in-line pasteurization is often applied, the microbiological status of the raw materials and of the processing facilities before pasteurization needs to be appropriate. Although vegetative microorganisms may be killed by pasteurization, products of their metabolism (which might not

be destroyed) remain. Intermediate product should be tested to verify that microbial (including ther- mophiles) growth does not occur in the aqueous phase. It is advisable to decontaminate any potentially suspect raw material, apply heat pasteurization to the aqueous phase or the complete emulsion and avoid recontamination by physical measures to effectively control infective pathogen hazards.

Composition. Reduced-fat spreads are mildly preserved products. Therefore, it is important to monitor all aspects of product composition that will affect their vulnerability toward microbiological

spoilage. The level of preservative, the equilibrium pH of the aqueous phase in the ﬁnished product, organic acids, and emulsion characteristics are all important properties that need to be veriﬁed.

Packaging. Because reduced-fat spreads are particularly mold-sensitive, adequate measures will speciﬁcally control mold contamination after the pasteurization stage and at packaging (incl. control of air and packaging material quality). Actually, proper hygiene during packaging will control spoilage in general, but the efﬁcacy of the control measures (incl. cleaning and disinfection of the packaging machine) must be monitored.

Distribution. Chilled distribution reduces the chance of mold problems during closed shelf-life and, in most cases, prevents the growth of other microbial contaminants as well. Refrigeration is certainly beneﬁcial during open shelf-life, where consumer use may cause contamination with for instance spoilage microorganisms. Low temperatures may also be required to maintain the desired physical properties of the product.

VI Butter

A Deﬁnitions Butter consists of at least 80% milk fat, a small amount of non-fat milk solids, water and, most often,

salt (sodium chloride). The fat phase is the continuous phase. A lactic starter culture, colors, and neutralizing agents may be used during manufacture. In the United States, Western-Europe and many other countries the composition of butter is strictly regulated: e.g. “the product with a milk fat content of not less than 80% but less than 90%, a maximum water content of 16%, and a maximum dry non-fat milk-material content of 2%” (EC, 1994). It is normally not permitted to add preservatives other than salt or anti-oxidants to butter (Murphy, 1990).

Consumers in Europe favor cultured butter, which is unsalted, or at most salted to a level not exceeding 0.5% (w/w). Sweet cream butter, often with 1.5–2% (w/w) salt, is sold almost exclusively in countries like UK, US, Australia, Japan, and India. Occasionally, consumers prefer unsalted, sweet butter. Cultured butter contains lactic acid instead of lactose; it may be of neutral pH, but usually has a

509 pH of 4.6–5.3. The primary ingredient for butter is cream, and the initial microﬂora of butter is therefore

OIL- AND FAT-BASED FOODS

to a large extent derived from the cream. In microbiological terms, unsalted, sweet butter is the most unstable type of butter.

B Important properties Like margarine, butter is a water-in-oil emulsion in which the water is ﬁnely dispersed (Keogh et al.,

1988; Madsen, 1990; van Zijl and Klapwijk, 2000). Different processing regimes may result in different microstructures. Microscopic methods have been developed to characterize the water dispersion in butter. Electron-microscopic examinations demonstrated that, in properly worked butter, the aqueous phase is almost exclusively present as isolated, globular, or elongated droplets with diameters <30 µm, covered with high-melting butter fat crystals (Buchheim and Dejmek, 1990). When the water in butter is compartmentalized in sufﬁciently small droplets that are well dispersed throughout the product, outgrowth of microorganisms will be limited and die-off may occur during storage. When the droplets are too coarse or when water channels have been formed, the aqueous phase composition (notably pH and salt-in-water concentration) becomes important for the keeping quality (Jensen et al., 1983).

C Methods of processing and preservation The processes for making butter have shown a shift away from batch to continuous processing (IDF,

1986b, 1987; Murphy, 1990, van Zijl and Klapwijk, 2000). The technology is quite advanced and cleanable-in-place, stainless steel equipment is common (IDF, 1996). Neutralization of cream is ac- complished in some countries by the addition of alkaline salts, like calcium oxide, sodium carbonate, or sodium hydroxide. Neutralization prevents excessive loss of fats during churning and eliminates objec- tionable oily or ﬁshy ﬂavors. This treatment has no bactericidal effect on the microbiological population of cream.

The butter process can be roughly divided into two stages: (1) a churning stage, in which the fat in the cream is concentrated to about 80% after which the so-called buttermilk is drained off, and (2) a working stage, in which the concentrated cream is worked into butter. In between, ingredients such as salt, colors, and concentrated starter cultures may be added.

The butter process starts with pasteurized cream. Pasteurization is designed to destroy relevant vegetative microorganisms, especially pathogens, but will not eradicate bacterial spore-formers and some of the more heat-resistant vegetative spoilage ﬂora. Pasteurization also aims at the destruction of enzymes in the raw cream that may reduce the organoleptic acceptability of the product and will completely liquefy the fat for subsequent control of crystallization (IDF, 1986b; Varnam and Sutherland, 1994). It is usual to pasteurize the cream after separation, even when the original milk is pasteurized (Murphy, 1990). Heat treatments (plate heat exchanger) involve temperatures in the range of 85–112 ◦ C (IDF, 1986b); 85–95 ◦

C for 10–30 s is most commonly used (Varnam and Sutherland, 1994). Batch pasteurization at longer holding times is obsolete or conﬁned to low technology plants in some areas of the world (van den Berg, 1988). Since there are no further decontamination treatments after the pasteurization step, subsequent processing should be designed to prevent the multiplication of those microorganisms surviving pasteurization and to minimize recontamination.

C, due to which the fat globules in the cream are converted to fat granules. The granules are then separated from the other constituents of the cream, leaving buttermilk. This results in a two-fold concentration of the fat in the cream. Most microorganisms are retained in the buttermilk, the aqueous phase in which the bacterial count is greater than that of the cream or the butter produced from it. Salt will be added, when at all, after the last point from which buttermilk drainage can occur. A concentration of 2% salt in product with 16% water

Churning of the cream involves intensive agitation at 5–7 ◦

MICROORGANISMS IN FOODS 6

will result in 11% salt in the aqueous phase [salt/(salt + water)]. This increased salt concentration may inhibit microorganisms, although microbial deterioration may occur even in highly salted butter when the salt is not uniformly distributed over the compartmentalized aqueous phase.

In mainland Europe, most butter traditionally has been made from soured cream using starter cultures and an aqueous phase pH of about 4.6. Since the resulting sour butter milk has limited market value, increasingly more use is being made of the so-called NIZO process which starts with sweet cream and yields sweet buttermilk. In this process, a (concentrated) lactic starter permeate (containing a high concentration of lactic acid) and, separately, an aromatic starter culture is worked directly into the butter after churning (Veringa et al., 1976; IDF, 1986b). These natural concentrates are commercially available. The resulting sweet buttermilk, after drying, has a higher market value than dried acid buttermilk. Butter resulting from this process has a desirable sour taste and aroma. It also has a reduced copper content and is therefore less sensitive to oxidation.

The butter is worked mechanically after churning to obtain the right physical properties, with the water being dispersed into minute droplets in a fat-continuous matrix. As with margarine, this greatly increases the shelf-stability of the product. A major process aim is to obtain small water droplets. Both under- working and over-working produce a ﬁnal butter with too coarse water droplet size distributions or even free moisture. As droplet size distributions are skewed with a tail of larger droplets, this means that about 50% of the total water volume should be in droplets with diameters <3 µm, while at the same time less than 5% of the water volume should be in droplets with diameters >10 µm (van Zijl and Klapwijk, 2000).

Butter may be packed in bulk (usually up to 25-kg units) or in consumer-size portions (10–500g). Parchment has traditionally been used as the primary packing material, but other materials have become available (IDF, 1987). Bulk-packed butter may be stored for long periods at very low temperatures, e.g. for 6 months at −15 ◦

C. Consumer-size portions should always be stored at chill temperatures (≤10 ◦

C or a year at −30 ◦

C) and even then have a storage time in general limited to 6–12 weeks. Re-packing of butter may result in enlargement of the water droplets, making the product microbiologically less stable.

D Microbial spoilage and pathogens Initial microﬂora. The microﬂora of butter is derived mainly from the cream used. The nature, sources,

and control of microorganisms likely to be present in industrial fresh milk and cream are discussed in Chapter 16. In the modern dairy situation, milk is collected on farms in refrigerated bulk tanks and transported by road tankers to the dairy factory. The microbiological quality of farm-separated cream differs considerably from that of fresh cream separated at a dairy plant, but farm-separation nowadays only occurs in areas with less advanced dairy industries. For instance, when cream is held on a farm for a week under poor hygienic conditions and without refrigeration, souring (e.g. by growth of Lactococcus lactis and other undesirable microorganisms) may have occurred, growth of yeasts and molds (e.g. Geotrichum candidum) may be abundant and Gram-negative aerobic bacteria (e.g. members of the genera Pseudomonas, Alcaligenes, Acinetobacter/Moraxella and Flavobacterium) may have proliferated, resulting in proteolytic and lipolytic changes (Foster et al., 1957).

Spoilage. Butter manufacture is a fairly sensitive process from a microbiological point of view and, therefore, microbial spoilage needs to receive due attention. Although farm-separated cream may be badly deteriorated, subsequent processing such as neutralization, vacuum treatment and the use of butter cultures may eliminate and/or mask off-ﬂavors. The butter from this cream is acceptable, although much inferior to that made from good-quality factory-separated cream.

Yeast and molds are important spoilage microorganisms of butter and can result in surface discol- oration and off-ﬂavor (references in Varnam and Sutherland, 1994; van Zijl and Klapwijk, 2000). A variety of mold genera (Penicillium, Oospora, Mucor, Geotrichum, Aspergillus and Cladosporium) have

511 been implicated. Some yeasts are lipolytic and can grow in the presence of high concentrations of salt,

OIL- AND FAT-BASED FOODS

at low pH, and at low temperatures. In retail samples of butter, Rhodotorula spp., Saccharomycopsis lipolytica, Cryptococcus laurentii, and Can. difﬂuens were reported as the predominant yeasts (Fleet and Mian, 1987).

Psychrotrophic Gram-negative bacteria such as Pseudomonas spp. and Flavobacterium spp. may develop and cause off-odor formation and rancidity (Driessen, 1983; Jooste et al., 1986; Champagne et al., 1993). Growth of Alteromonas putrefaciens or Flavobacterium malodoris may lead to surface taints (Foster et al., 1957; Jooste et al., 1986) very quickly affecting the mass of the product and accompanied by the development of a putrid, decomposed, or cheesy flavor (apparently from isovaleric acid or a closely related compound). Certain Pseudomonas spp. are associated with the formation fruity odors or black discolorations of the butter (Foster et al., 1957). A variant of Lactococcus lactis (formerly Streptococcus lactis var. maltigenes) may cause the so-called malty-flavor defect related to the formation of 3-methylbutanal (Jackson and Morgan, 1954). When the organism grows extensively in cream before pasteurization, the flavor can carry over to the pasteurized cream.

Rancid ﬂavor comes mainly from free butyric acid that arises from the hydrolysis of butter fat. This reaction may be catalyzed by naturally occurring lipase in the milk or by enzymes secreted by microorganisms. Milk lipase is destroyed by pasteurization (Driessen, 1983). Most, but not necessarily all, lipases of microbial origin will be fully inactivated by pasteurization. The carry-over of residual enzyme activity, bacterial toxins, and off-ﬂavor to the pasteurized cream are the major reasons to strictly limit microbial growth in non-pasteurized cream, even though the previously mentioned microorganisms will all be destroyed by proper pasteurization.

Pathogens. Since the milk used for the production of butter may carry vegetative pathogens (e.g. L. monocytogenes, E. coli O157:H7; Chapter 16), pasteurization of the milk upon arrival at the dairy factory and measures to control of post-pasteurization recontamination are important for microbiological safety (Murphy, 1990; van Zijl and Klapwijk, 2000). Essentially all commercial butter is made from pasteurized cream and since the physical–chemical characteristics of butter (notably the ﬁne water dispersion and fat continuity) are more inhibitory than those of pasteurized milk, butter should present even less of a food-borne disease problem than pasteurized milk. Sims et al. (1969) reported that butter made from inoculated cream supported growth of Salmonella at 25 ◦

C, while populations decreased during storage below 4.4 ◦

C. El-Gazzar and Marth (1992) reported on large decreases in Salmonella for unsalted butter stored at −17.8 ◦

C. Lanciotti et al. (1992) observed growth of L. monocytogenes in light butter and concluded that the pathogen might be less affected by space and the nutritional lim- itations of the compartmentalized structure compared with butter. Holliday et al. (2003) investigated the survival and growth characteristics of mixtures of 5 serotypes of Salmonella, ﬁve strains of E. coli O157:H 7, and 6 strains of L. monocytogenes in 3 types of commercial butter: sweet cream whipped salted butter (pH 6.4; 78% fat), sweet cream whipped unsalted butter (pH 4.51; 78% fat), and salted light butter (pH 4.58; 43% fat; with preservatives). The products were subjected to temperature abuse, by holding at

C and 23.3 ◦

C under high relative humidity (85%) for 1 h to induce condensation of water on the surface, before

storing at 4.4 ◦ C or 21

C for up to 21 days. Sweat cream whipped salted butter supported surface growth of all three pathogens and of only L. monocytogenes during storage at 21 ◦

C, respectively. The other two products did not support growth of any of the three pathogens at either temperature for the duration of storage. All pathogens tested were inactivated more rapidly in products stored at 21 ◦

C and at 4.4 ◦

C than at 4.4 ◦

C, and in products containing preservatives and acidulants. It is advisable to conduct challenge studies to determine pathogen survival and growth in butter, especially under abuse conditions, and assure that product characteristics include additional hurdles to pathogen growth. A known antimicro- bial such as garlic might be considered to provide an additional defence to growth of certain pathogens (Zhao et al., 1990), although an outbreak of Campylobacter enteritidis involving garlic butter has been

MICROORGANISMS IN FOODS 6

reported (Anonymous, 1996). Adler and Beuchat (2002) investigated the viability of mixtures of different sero-types and strains of three pathogens (5 serotypes of Salmonella, ﬁve strains of E. coli O157:H7 and 6 strains of L. monocytogenes) inoculated onto unsalted butter with or without garlic in dependence of the storage temperature. None of the pathogens was able to grow, although they all retained viability at

C regardless of the presence of garlic; addition of garlic speeded-up inactivation at 21 ◦

C and 37 ◦ C.

A number of cases concerned contamination with Staph. aureus or its toxin, but over the years

a variety of vegetative pathogenic bacteria were implicated as well. Nevertheless, considering the extremely large volumes that butter is marketed on, there are relatively few reported cases of butter associated food-borne illness to date.

In 1970, butter was voluntarily recalled by a manufacturer in the United States because of excessively high numbers of bacteria and the suspicion of staphylococcal contamination (Anonymous, 1970). In the same year, a case of food poisoning occurred in a restaurant implicating whipped butter prepared from butter contaminated by Staph. aureus (US-DHEW, 1970). Notably, butter from the same brand was implicated as the cause of a single case of typical staphylococcal food poisoning and was shown to contain staphylococcal enterotoxin A. (US-DHEW, 1970). In 1977, whipped butter produced by a single manufacturing plant in the United States was implicated in a multi-state outbreak of presumed

staphylococcal food poisoning, which involved over 100 people; up to 10 7 cfu/g Staph. aureus were isolated from lots of the whipped butter involved (US-DHEW, 1977). Whipping of butter increases the risk of Staph. aureus growth, because water or milk may be added to the butter before whipping and because the salt level in the ﬁnal product is reduced. Therefore, the product should be subjected to strict hygiene, refrigerated storage (≤7 ◦

C) and a restricted shelf-life. The toxin of Staph. aureus is very heat stable (Brunner et al., 1991) and, once formed as a result of poor sanitary conditions in the pre-pasteurization stage, might carry over to the pasteurized product.

A food poisoning caused by consumption of blended margarine and butter products contaminated by Staph. intermedius occurred in the United States in 1991 and involved over 265 people; since a single strain was identified as the causative agent, a common source of contamination rather than post- process contamination was suggested (Anonymous 1992b; Khambaty et al., 1994; Bennet, 1996). From Germany, an incident with green butter contaminated with Citrobacter freundii was reported; the butter was made by adding organically grown parsley and a genetically identical Cit. freundii was isolated from most patients and from the parsley of the organic garden (Tschäpe et al., 1995). An outbreak in 1995 of C. enteritis involved about 30 people who had eaten at a restaurant in Louisiana and appeared to be associated with garlic butter prepared by the restaurant (Anonymous, 1996). In several cases, L. monocytogenes (Marth and Ryser, 1990; Massa et al., 1990; Harvey and Gilmore, 1992) or Salmonella spp. (Cavalcante dos Santos et al., 1995) contamination was suspected, although these bacteria were not detected in the samples studied. In a case-control study of a cluster of peri-natal listeriosis cases in California carried out over a period of 6 months in 1987–1988, butter was identified as a possible vehicle for infection (Chun et al., 1990) although no direct evidence was obtained. A recall in the United States in 1994 involved unsalted and lightly salted butter contaminated with L. monocytogenes (Anonymous, 1994; Proctor et al., 1995), but no cases of food poisoning were linked to this contamination.

A serious outbreak of food poisoning occurred in 1999 and implicated butter from a Finnish dairy plant. Butter in small-sized packages (7 and 10 g) were found to be contaminated with L. monocytogenes serotype 3A (Lyytik¨ainen et al., 1999, 2000). Poor sanitation within the plant led to contamination of the butter packages. Eighteen people developed listeriosis, four of whom died. The mean age of patients was 57 years (range 18–85); all had serious underlying illnesses and were undergoing treatment in several hospitals. Most of the disease-causing strains (14/18) were of a rare serotype 3a which was isolated from samples taken from different places in the butter production facility, including environmental and butter samples from the dairy plant and butter samples from the cold store of the dairy. Isolates were also obtained from the kitchen of a hospital. The isolates from butter and epidemic isolates were

513 indistinguishable by pulsed-ﬁeld gel electrophoresis (PFGE) and their PFGE-type had not been isolated

OIL- AND FAT-BASED FOODS

previously from any other foods in Finland. L. monocytogenes was detected at low levels (<10 2 cfu/g) in most of the small butter packs examined. One pack contained >10 4 cfu/g, a level in food well documented to cause outbreaks of listeriosis. Estimates of the doses of the pathogen actually ingested range from 14– 20 cfu/day and 2.2 × 10 4 and 3.1 × 10 5 , based on hospital kitchen data and contamination found in retail samples, respectively (Maijala et al., 2001). L. monocytogenes may survive for months in butter when temperatures are very low (−18 ◦

C (Olsen et al., 1988). European Union Directives specify that L. monocytogenes should not be detectable in 1 g of butter.

C) and will grow slowly if temperatures are 4 ◦

C or 13 ◦

E CONTROL (reduced-fat spread)