Strains of penicillium camemberti

Abstract

The present invention relates to strains of Penicillium camemberti and to the use thereof for the preparation of food products, for example of dairy and/or vegetable origin, such as the ripening of soft cheeses having a moldy and/or mixed crust, in particular camembert.

Claims

1. An isolated strain of Penicillium camemberti having a lag time on a milk culture medium at 4° C. longer than 19 days, said strain being strain I-5305, registered with the CNCM on 5 Apr. 2018.

2. A method for preparing a food product comprising a step of fermenting a food product with the Penicillium camemberti strain according to claim 1 thereby producing said food product.

3. The method according to claim 2, wherein the food product is a cheese, a vegetable matrix or a mixture of a vegetable matrix and a dairy material.

4. The method according to claim 2 wherein the fermentation step is performed with a mixture of surface strains comprising the Penicillium camemberti strain having a lag time on milk culture medium at 4° C. longer than 19 days, said strain being strain I-5305, registered with the CNCM on 5 Apr. 2018 and at least one other surface strain.

5. The method according to claim 4, wherein said other surface strain is Geotrichum candidum.

6. A fermented food product obtained by the method according to claim 2.

Description

FIGURES

(1) FIG. 1 pH kinetics in the interior and rind of cheeses, made with strain I-5305 and PC 12 (strain CHOOZIT PC12, Danisco).

(2) FIG. 2 Kinetics of the residual concentration of lactate in the interior and rind of cheeses, made with strain I-5305 and PC 12 (strain CHOOZIT PC12, Danisco).

EXAMPLE 1—CHARACTERISATION OF THE FUNCTIONAL PROPERTIES OF P. CAMEMBERTI STRAINS OF THE INVENTION

(3) Materials and Methods:

(4) a) Strains studied:

(5) The strains studied are the following: I-5302, I-5304, I-5305, I-5307, I-5308, I-5309, I-5310, I-5311 are kept frozen at −80° C., in the form of spores suspended in a 15% glycerol cryoprotectant. The spores are obtained after culture on a synthetic medium, solid medium of the PDA type (Potato Dextrose Agar marketed by Biokar), PC 12 and PC SAM3 (supplier: Danisco) are marketed in liquid form. They are kept frozen (at −80° C.) in a mixture consisting of 50% liquid ferment and 50% cryoprotectant at 30% glycerol, PCTN, PC PR1, PC TAM5 (supplier: Lallemand) and PC A1 swing (supplier: CHR HANSEN) are marketed in lyophilised form. They are kept frozen at −80° C., according to the following mixture 2 g lyophilisate+18 g cryoprotectant at 15% glycerol,

(6) b) Medium used:

(7) The medium used is a dairy medium, the dairy and acidified MRP.

(8) Media: Dairy model (MRP)

(9) TABLE-US-00001 TABLE 1 Composition of MRP medium for 1 liter g Milk base 727 Type E bacteriological agar 18.2 90% pure lactic acid L 9.1 50% sodium lactate 9 Casein acid hydrolysate 0.09 Sodium chloride 11.8

(10) The milk base is composed of 11.22% standard anhydrous milk fat, 11.48% ultra-filtered skimmed milk powder and 77.30% water.

(11) c) Inoculation of the containers:

(12) The dilutions are adjusted according to the cell count of the different ferments, in order to obtain isolated colonies and thus measure their diameter more easily.

(13) d) Observations:

(14) The containers are observed every day from D+3.

(15) e) Observed parameters:

(16) The growth of the strains is studied by measuring the radial growth (which enables the calculation of the recovery rate) and their lag time. Measuring the recovery rate:

(17) This is obtained by the daily measurement of the diameter of the colonies.

(18) The recovery rate in cm per day is defined by the slope of the straight line obtained from the graph: diameter of colonies (cm) as a function of time (days). Measure of the lag time:

(19) Its calculation is made with respect to the equation of the line representing radial growth.

(20) The strains tested have different physiological states which can be in liquid form, in the form of lyophilsate or which have been grown on agar medium.

(21) Two series of characterisation were therefore launched one week apart, while respecting the day of inoculation to have identical reading days for both series.

(22) Results: The lag time:

(23) TABLE-US-00002 TABLE 2 Measure of lag time on MRP medium at 4° C. Strain Lag time (days) PC PR1 16.26 PC 12 16.59 PC TAM5 16.91 PC TN 17.82 PC A1 swing 18.33 PC SAM3 18.58 I-5311 19.53 I-5304 19.55 I-5307 20.04 I-5308 21.14 I-5305 21.23 I-5302 21.28 I-5309 22.23 I-5310 36.25 The recovery rate:

(24) TABLE-US-00003 TABLE 3 Measure of recovery rate on MRP medium at 4° C. Strain Recovery rate (cm/day) I-5305 0.04 I-5311 0.12 I-5310 0.14 PC TN 0.16 PC PR1 0.16 PC TAM5 0.16 PC 12 0.16 I-5307 0.17 PC SAM3 0.17 PC A1 swing 0.17 I-5304 0.19 I-5309 0.19 I-5308 0.22 I-5302 0.23

(25) Results:

(26) The commercial strains have a recovery rate of between 0.16 and 0.23 cm/day and a lag time of between 16 and 18 days.

(27) The strains according to the invention (I-5311, I-5304, I-5307, I-5308, I-5305, I-5302, I-5309 and I-5310) have a longer lag time, systematically longer than 19 days on MRP medium at 4° C.

(28) Furthermore, some of them (I-5305, I-5310 and I-5311) have a recovery rate of less than 0.15 cm/day, on MRP medium at 4° C.

(29) The properties of the P. camemberti strains according to the invention make them excellent candidates for use for the slow ripening of cheeses with a bloomy and/or mixed rind, thus allowing the longer preservation of these cheeses.

EXAMPLE 2—USE OF DIFFERENT STRAINS OF P. CAMEMBERTI FOR THE PRODUCTION OF SOFT CHEESE WITH A BLOOMY RIND FROM A LACTIC CURD ENRICHED WITH FAT IN A VAT

(30) Materials and Methods:

(31) a) Description of the steps of production:

(32) The method for preparing a soft cheese with a bloomy rind from a lactic curd enriched with fat in a vat comprises the following main steps: cow's milk enriched by the addition of whole cream or retentate having the characteristics of a target of 30±5 g/l protein matter (PM) and 110±10 g/l fat (F), is pasteurised according to the standard procedures of a person skilled in the art, commercial mesophilic lactic ferments (2 and 10 g/l, such as for example a mixture of Lactococcus lactis ssp cremoris, Lactococcus lactis ssp lactis, Lactoccoccus lactis ssp lactis biovar diacetylactis, Leuconostoc) and ripening (comprising yeasts and moulds) are then added in the usual doses to make a lactic curd, the milk is renneted at a pH equal to or less than 6.50 in vats of 200 I-500 I in suitable doses and the milk is left to curdle, the forming is performed at a pH less than or equal to 4.90, the product is then left to rest before being removed from the moulds by turning over if necessary, the salting is carried out dry, the ripening step is performed before proceeding to packaging.

(33) The P. camemberti strains can be inoculated either directly with lactic ferments and other ferments at doses between 0.1 to 2 doses per 100 I of milk (1 dose=2.10.sup.9 spores/ml) or as a spray after salting at a dose between 0.4 and 5 doses/I of water.

(34) b) Studied strains:

(35) Strain I-5305 as well as strains VELV-TOP®PC 1131 of Lallemand and VELV-TOP®PC PR1 of Lallemand were each used for producing a cheese at the same doses and in the same conditions.

(36) Organoleptic Results:

(37) The soft cheeses with a bloomy rind produced with each of these strains were submitted to a panel of experts for an evaluation of the products after packaging at an optimum age (25-30 days) with regard to 4 criteria: external appearance, ease of cutting, texture and taste.

(38) Table 4 below summarises the main advantages observed between the product made from strain I-5305 compared to products obtained with strain IB1 and with strain PR1. These comments are from two independent replicates.

(39) TABLE-US-00004 TABLE 4 The main advantages observed between the product made with ferment I- 5305 relative to the product made with ferment IB1 and ferment PR1. Advantages I-5305 versus Advantages I-5305 versus IB1 PR1 External appearance Even surface, no hollows Ridges of product less marked Ease of cutting and Rind slightly less thick, less The rind is integral to the appearance when cut crisp interior The rind is integral to the The interior is less fluid interior Texture Good when cut, less soft Rind less perceptible in mouth Taste Less mature, less earthy, Less buttery less bitter, more creamy, slight fungus *The date of inoculation is considered to be D + 1.

(40) The cheese obtained with strain I-5305 makes it possible to obtain a product after packaging which is significantly whiter with a smoother and more homogenous surface; it has good cutting behaviour with a rind integral to the soft cheese, with a marked creamy taste with a typical fungus note with no defects at optimum tasting age, such as bitterness and earthiness.

(41) The cheese obtained with strain I-5305 is judged, at the same age, to have a longer shelf life according to the established criteria of colour, lack of bitterness and smoothness below the rind which are desirable in a mature product.

EXAMPLE 3—USE OF DIFFERENT STRAINS OF P. CAMEMBERTI FOR PRODUCING A GOAT'S CHEESE IN VAT TECHNOLOGY

(42) Materials and Methods:

(43) a) Description of production steps:

(44) The method of preparing a goat's cheese with vat technology with a 100% P. camemberti rind comprises the following main steps: a goat's cheese enriched by the addition of cream then delactosed to 17% having the characteristics or a target of 33±2 g/l protein matter and 46±2 g/l fat, is pasteurised according to the standard methods used by a person skilled in the art, commercial mesophilic lactic ferments (for example a mixture of Lactococcus lactis ssp cremoris, Lactococcus lactis ssp lactis, Lactoccoccus lactis ssp lactis biovar diacetylactis, Leuconostoc at 3 g for 100 L) can be inoculated to enable cold maturation, from 0.15 to 0.2 pH unit, the material is then heat treated and preserved. thermophilic lactic cultures are added (Streptococcus thermophilus at 4 g/100 l), the renneting of the milk is carried out at a pH equal to or lower than 6.35 in 200 I-500 I vats in suitable doses and left under serum, the forming is carried out at a pH lower than or equal to 6.40, the product is then left to rest before being demoulded by turning over as necessary at a pH of 5, the salting is carried out dry, the ripening step is performed before proceeding to packaging.

(45) The P. camemberti strains can be inoculated either directly with the lactic ferments and other ferments at doses between 0.4 dose for 100 I milk or by spraying for 24 h after salting at 1 dose/l of water.

(46) b) Studied strains:

(47) Strain I-5305 as well as strains PC TN (VELV-TOP PC TN, Lallemand) and PC 12 (Danisco) are tested at the same doses and in the same conditions.

(48) Organoleptic Results:

(49) The soft cheeses with bloomy rind produced were submitted to a panel of experts for evaluation within the optical age window of D+28.

(50) Table 5 below shows the main characteristics of cheeses inoculated with 50% PC 12 and PC TN and those having 100% of strain I-5305.

(51) TABLE-US-00005 TABLE 5 Main characteristics of cheeses inoculated with 50% PC 12 and 50% PC TN and one having 100% CNCM I-5305 tasted by a panel of experts on D + 28. Composition of the inoculation of the rind 50% PC 12 and 50% PC TN 100% I-5305 Appearance White fluffy fairly Fluffy white but to a lesser extensive extent, less relief, more level Smell Weak smell - fungus Weak smell - more cave Texture Very medium - Texture + melt - rind not very marked a little firmer - rind not very marked Taste Fungus taste very present Fungus taste present- no Slight bitterness to finish bitterness Desirable characteristics of products marked in bold.

(52) From a semi-ripened stage, there is clearly a qualitative advantage of the series of cheeses produced with I-5305 in all of the criteria required during the production of quality cheese with a bloomy rind.

EXAMPLE 4—USE OF DIFFERENT P. CAMEMBERTI STRAINS FOR PRODUCING LOW-FAT SOFT CHEESE WITH A BLOOMY RIND ACCORDING TO A METHOD OF ULTRAFILTRATION

(53) Materials and Methods:

(54) a) Description of production steps:

(55) The method of preparation of a low-fat soft cheese with a bloomy rind according to a method of ultrafiltration comprises the following main steps: preparation of the precheese which comprises the retentate, the homogenised cream, the salt in the mass, the gelatine, the betacarotene, dosage and incorporation of mesophilic lactic ferments, P. camemberti and rennet, acidification at a pH lower than or equal to 5.35, cooling, demoulding and brining, drying, ripening, drying before packaging, packaging, storage.

(56) b) Studied strains:

(57) Strain I-5305 as well as strain PC 12 (strain CHOOZIT PC 12, Danisco) are tested at the same doses and in the same conditions.

(58) Results:

(59) a) Analytic results:

(60) The pH kinetics carried out in comparison on the two cheeses reveals that the cheese made solely with strain PC 12 (CHOOZIT PC 12, Danisco) has a greater rise in pH both in the interior and in the rind by a delta of one pH unit at 50 days (FIG. 1).

(61) From 30 days after production, the kinetics of residual lactate concentration within the two cheeses begin to diverge with a higher residual concentration on the rind and in the interior for the cheese made with strain I-5305. 50 days after manufacture, an evolution towards a stabilised core-rind equilibrium around a lactate concentration of 4 g/kg is observed for strain I-5305 in comparison with a much lower residual concentration for the cheese with strain PC 12 (CHOOZIT PC12, Danisco). The residual presence of lactate in the rind, on the optimum use-by date, makes it possible for the Penicillium camemberti strain to continue to consume lactate instead of proteins, which gives it a better taste on the optimum use-by date.

(62) These results show significantly a cheese with a lower pH with rind when combined with strain I-5305. This more acidic pH limits the formation of an insoluble calcium layer that causes the formation of a granular rind, which is an organoleptic defect in soft cheese with a light bloomy rind obtained by an ultrafiltration process.

(63) b) Organoleptic results:

(64) The soft cheeses with a bloomy rind were submitted to a panel of experts for evaluation of the products at different stages of their life, to assess the evolutionary aspect of the product with regard to different criteria: external appearance, rind quality and taste.

(65) Table 6 below summarises the main differences observed between the product made with strain CNCM I-5305 compared to the product made with strain PC 12 (CHOOZIT PC12, Danisco).

(66) TABLE-US-00006 TABLE 6 Main differences observed between the product made from ferment I-5305 compared to the product made with ferment PC12 Semi-mature Mature product at Young product product optimal use-by date (D − 21)* (D + 29) (D + 50) External Product much appearance whiter Quality The cheese The cheese The cheese of the rind obtained with obtained with obtained with I- I-5305 has a I-5305 has a 5305 has a less softer rind, less grainy rind detached, thicker, more runny, drier rind slightly grainy, less detached Taste The cheese The cheese obtained with obtained with I-5305 I-5305 is more has a less acidic, less pronounced mould ripened taste *The inoculation date is considered as D + 1.

(67) These results show that, at an equivalent age, the benefits of cheese made with strain I-5305 may represent an improvement in quality during the period of the optimal use-by date.

EXAMPLE 5—USE OF A MIXTURE OF DIFFERENT P. CAMEMBERTI STRAINS FOR PRODUCING SOFT CHEESE WITH A BLOOMY RIND ACCORDING TO AN ULTRAFILTRATION METHOD

(68) Materials and Methods:

(69) a) Description of production steps:

(70) The method for preparing low-fat soft cheese with a bloomy rind according to an ultrafiltration method in combination with other P. camemberti strains comprises the following main steps: preparing the precheese which comprises the retentate, the fermented milk, the cream, mixing with the preparation comprising added dairy proteins and flavouring, homogenising with degassing then heat treatment, incorporating lactic ferments (Lactococcus lactis ssp lactis biovar diacetylactis, Streptoccocus thermophilus, Lactobacillus delbrueckii ssp bulgaricus), of P. camemberti and rennet, dosing and acidification at a pH higher than or equal to 4.85, cooling, demoulding and brining, drying, ripening, drying, the ripening step is performed before proceeding to packaging, storage.

(71) b) Studied strains:

(72) Strain I-5305 as well as strain PC 12 (strain CHOOZIT PC12, Danisco) are tested with 2 final doses (D) of Penicillium camemberti for 100 kg retentate with ratios of PC12/I-5305 ranging from 0.5 to 1.5 D (1D=2.10.sup.9 spores/mL).

(73) Organoleptic Results:

(74) The soft cheese with bloomy rind produced were submitted to a panel of experts throughout their life cycle

(75) Table 7 below shows the main characteristics of cheeses inoculated with 100% of strain PC 12 (strain CHOOZIT PC12, Danisco) and those having a combination of 50% strain of I-5305 associated with 50% of strain PC 12 (strain CHOOZIT PC12, Danisco).

(76) TABLE-US-00007 TABLE 7 Main observations between the product made from strain PC 12 (strain CHOOZIT PC 12, Danisco) alone or in combination with strain I-5305 and monitoring of their evolution during ripening at D + 31, D + 38, D + 45, D + 59). Composition of the inoculation of the rind D + 31 D + 38 D + 45 D + 59 100% PC 12 Rind White Slightly yellow Yellow slightly thick appearance, appearance appearance, taste of rind intensification “moisi-sylv{acute over (e)}- of fungus” “moisi-sylvé” sandy rind note texture grainy rind 50% I-5305 White White White White 50% PC 12 appearance, appearance, appearance, appearance, below white below white below white below white rind rind rind rind *The date of inoculation is considered as D + 1 Underlined: appearance of ripening markers per product.

(77) At the semi-ripening stage of production (D+38 and D+45), there were significant differences between the 2 products with changes in appearance, rind texture and taste revealing a more advanced ripening stage on the cheese rind with the 100% strain PC 12 (strain CHOOZIT PC12, Danisco). The cheese with the addition of strain I-5305 allows the product to be stabilised in terms of colour and taste up to the product's use-by date, while maintaining a superior product quality throughout its shelf life.

EXAMPLE 6—USE OF A MIXTURE OF DIFFERENT STRAINS OF P. CAMEMBERTI FOR THE PRODUCTION OF GOAT'S CHEESE ACCORDING TO AN ULTRAFILTRATION METHOD, IN MIXED RIND

(78) Materials and Methods:

(79) a) Description of production steps:

(80) The method of preparation of a goat's cheese according to an ultrafiltration method, in mixed rind comprises as the following main steps: preparing the pre-cheese which comprises the retentate, the cream, dairy proteins, heat treatment, incorporating lactic strains (Lactococcus lactis ssp lactis, Lactococcus lactis ssp cremoris, Streptococcus thermophilus, Lactobacillus delbrueckii ssp bulgaricus) and ripening ferments (P. camemberti and Geotrichum candidum) and rennet, dosage and acidification, up to a target pH of 5, cooling, demoulding and brining, ripening, drying, packaging, storage.

(81) b) Studied strains:

(82) Strain I-5305 as well as strain VELV-TOP®PC PR1 of Lallemand were tested at a dose of 4 doses/100 kg. They can be applied in combination with strains of Geotrichum candidum at 0.2 dose/100 kg (1 dose=8.10.sup.7 UFC/ml).

(83) Organoleptic Results:

(84) The soft cheeses produced were submitted to a panel of experts for evaluation in the optimum age window at D+29.

(85) Table 8 below shows the main characteristics of cheeses according to the ripening strategy used.

(86) TABLE-US-00008 TABLE 8 Comparison of cheeses inoculated with PC PR1 and I-5305 mixed with two ferments of Geotrichum candidum tasted by a panel of experts at D + 28. Geotrichum Geotrichum Geotrichum Geotrichum candidum and candidum and candidum and candidum and PC PR1 I-5305 PC PR1 I-5305 Appearance Crinkled Crinkled Folded flower, Folded flower, flower, yellow flower, very yellow side very white and side and white and not and bottom & not humid bottom & humid humid to humid to touch touch Cut Fine rind Very fine rind Fine rind Very fine rind Texture Sticky but Rind present, Sticky smooth remains crisp Rind barely smooth perceptible Rind barely perceptible Taste/smell Intense cave Taste: softer, Smell: very Taste: softer, type smell rounder, less intense rounder, less aggressive aggressive Smell: less Smell: more cave fungus In bold: the desired characteristics of the products.

(87) It appears that all of the trials for mixed rind with the I-5305 strain show a qualitative gain in terms of colour, quality of the rind which is fine and barely perceptible and with a weaker smell.