FNS  Vol.4 No.9 B , September 2013
Selective Lactococcus Enumeration in Raw Milk
Abstract: The Lactococcus diversity in cow and goat raw milk was investigated. To do so, a protocol had to be established for the specific enumeration of lactococci. Eight agar media and one control medium were analysed to compare their proficiency in evaluating the Lactococcus population in raw milk: M17 Nal, Elliker, modified Elliker, PCA + milk, modified KCA, modified Chalmers, Turner, FSDA. The M17 medium was used as reference. Eighteen pure strains were tested on these media for their selectivity towards lactococci: six Lactococcus species or subspecies, three Leuconostoc, three Enterococcus, two Lactobacillus, one Streptococcus thermophilus, one Pseudomonas fluorescens, one Escherichia coli and one Staphylococcus aureus. All these bacteria were chosen for their regular presence in raw milk. The KCA medium proved to be the most selective towards lactococci, on condition that 1) we discriminated the colonies using the catalase test and 2) we subtracted the Enterococcus population counted on BEA. However, it was not possible to separate the Streptococcus from the Lactococcus colonies on KCA. The “Lactococcus-like” population including these two genera was estimated at a mean level of 3.18 log(cfu)/mL and 4.14 log(cfu)/mL in cow and goat raw milk respectively. This is consistent with the data already published.
Cite this paper: L. Gemelas, V. Rigobello, M. Ly-Chatain and Y. Demarigny, "Selective Lactococcus Enumeration in Raw Milk," Food and Nutrition Sciences, Vol. 4 No. 9, 2013, pp. 49-58. doi: 10.4236/fns.2013.49A2008.

[1]   L. Quigley, O. O’Sullivan, T. P. Beresford, R. P. Ross, G. F. Fitzgerald and P. D. Cotter, “Molecular Approaches to Analysing the Microbial Composition of Raw Milk and Raw Milk Cheese,” International Journal of Food Microbiology, Vol. 150, No. 2-3, 2011, pp. 81-94. doi:10.1016/j.ijfoodmicro.2011.08.001

[2]   M. Vacheyrou, A.-C. Normand, P. Guyot, C. Cassagne, R. Piarroux and Y. Bouton, “Cultivable Microbial Communities in Raw Cow Milk and Potential Transfers from Stables of Sixteen French Farms,” International Journal of Food Microbiology, Vol. 146, No. 3, 2011, pp. 253-262. ,

[3]   S. Buchin, V. Delague, G. Duboz, J. L. Berdague, E. Beuvier, S. Pochet and R. Grappin, “Influence of Pasteurization and Fat Composition of Milk on the Volatile Compounds and Flavor Characteristics of a Semi-Hard Cheese,” Journal of Dairy Science, Vol. 81, No. 12, 1997, pp. 3097-3108. ,

[4]   E. Beuvier, K. Berthaud, S. Cegarra, A. Dasen, S. Pochet, S. Buchin and G. Duboza, “Ripening and Quality of Swiss-Type Cheese Made from Raw, Pasteurized or Microfiltered Milk,” International Dairy Journal, Vol. 7, No. 5, 1997, pp. 311-323. doi:10.1016/S0958-6946(97)00015-0

[5]   Y. Bouton, P. Guyot and R. Grappin, “Preliminary Characterization of Microflora of Comté Cheese,” Journal of Applied Microbiology, Vol. 85, No. 1, 1998, pp. 123-131. ,

[6]   Y. Demarigny, E. Beuvier, S. Buchin, S. Pochet and R Grappin, “Influence of Raw Milk Microflora on the Characteristics of Swiss-Type Cheeses: II. Biochemical and Sensory Characteristics,” Le Lait, Vol. 77, No. 1, 1997, pp. 151-167. doi:10.1051/lait:1997110

[7]   N. Desmasures, “Etude de laits de Haute Qualité: Caractéristiques Microbiologiques et Aptitude à la Transformation en Camembert au lait cru,” Ph.D. Dissertation, University of Caen, Caen, 1995.

[8]   E. Beuvier and S. Buchin, “Raw Milk Cheeses,” In: P. F. Fox, P. L. H. McSweeney, T. M. Cogan and T. P. Guinee, Eds., Cheese: Chemistry, Physics and Microbiology, 3rd Edition, Tec & Doc Editions, Vol. 1, Paris, 2004, pp. 319345. doi:10.1016/S1874-558X(04)80072-1

[9]   E. Casalta, R. Zennaro, M.-X. Maroselli and R. Legouar, “Effect of Specific Starters on Microbiological, Biochemical and Sensory Characteristics of Venaco, a Corsican Soft Cheese,” Sciences de Aliments, Vol. 17, No. 1, 1997, pp. 79-94.

[10]   B. E. Terzaghi and W. E. Sandine, “Improved Medium for Lactic Streptococci and Their bacteriophages,” Applied and Environmental Microbiology, Vol. 29, No. 6, 1975, pp. 807-813.

[11]   N. Turner, W. E. Sandine, P. R. Elliker and E. A. Day, “Use of Tetrazolium Dyes in an Agar Medium for Differentiation of Streptococcus lactis and Streptococcus cremoris,” Journal of Dairy Science, Vol. 46, No. 5, 1963, pp. 380-385. doi:10.3168/jds.S0022-0302(63)89059-1

[12]   G. Waes, “The Enumeration of Aromabacteria in BD Starters,” Netherlands Milk and Dairy Journal, Vol. 22, 1968, pp. 29-39.

[13]   A. M. Huggins and W. E. Sandine, “Differenciation of Fast and Slow Milk Coagulating Isolates in Strains of Streptococci,” Journal of Dairy Science, Vol. 67, No. 8, 1984, pp. 1674-1679. doi:10.3168/jds.S0022-0302(84)81491-5

[14]   N. Desmasures and M. Gueguen, “Monitoring the Microbiology of High Quality Milk by Monthly Sampling over 2 Years,” Journal of Dairy Research, Vol. 64, No. 2, 1997, pp. 271-280. ,

[15]   J. F. Chamba, G. Bonnaz and P. Bourg, “Comparaisons de Diverses Méthodes de Dénombrement de la Flore Acidifiante du lait cru,” Le Lait, Vol. 61, No. 609-610, 1981, pp. 555-567. ,

[16]   V. Vanos and L. Cox, “Rapid Routine Method for the Detection of Lactic Acid Bacteria among Competitive Flora,” Food Microbiology, Vol. 3, No. 3, 1986, pp. 223234. doi:10.1016/0740-0020(86)90003-1

[17]   M. Dalmasso, C. Hennequin, D. Duc and Y. Demarigny, “Influence of Backslopping on the Acidifications Curves of ‘Tomme’ Type Cheeses Made during 10 Successive Days,” Journal of Food Engineering, Vol. 92, No. 1, 2009, pp. 50-55. doi:10.1016/j.jfoodeng.2008.10.019

[18]   P. R. Elliker, A. W. Anderson and G. Hannesson, “An Agar Culture Medium for Lactic Acid Streptococci and Lactobacilli,” Journal of Dairy Science, Vol. 39, No. 11, 1956, pp. 1611-1612. ,

[19]   Z. Y. Pu, M. Dobos, G. K. Y. Limsowtin and I. B. Powell, “Integrated Polymerase Chain Reaction-Based Procedures for the Detection and Identification of Species and Subspecies of the Gram-Positive Bacterial Genus Lactococcus,” Journal of Applied Microbiology, Vol. 93, No. 2, 2002, pp. 353-361. ,

[20]   V. Michel, A. Hauwuy and J. F. Chamba, “La Flore Microbienne de laits Crus Vache: Diversité et Influence des Conditions de Production,” Le Lait, Vol. 81, No. 5, 2001, pp. 575-592. doi:10.1051/lait:2001151

[21]   D. Corroler, I. Mangin, N. Desmasures and M. Gueguen, “An Ecological Study of Lactococci Isolated from Raw Milk in the Camembert Cheese Registered Designation of Origin Area,” Applied and Environmental Microbiology, Vol. 64, No. 2, 1998, pp. 4729-4735.

[22]   Y. Demarigny, C. Sabatier, N. Laurent, S. Prestoz, V. Rigobello and M. J. Blachier, “Microbiological Diversity in Natural Whey Starters Used to Make Traditional Rocamadour Goat Cheese and Possible Relationships with Its Bitterness,” Italian Journal of Food Science, Vol. 18. No. 3, 2006, pp. 251-266.

[23]   F. Grattepanche, P. Audet and C. Lacroix, “Milk Fermentation by Functional Mixed Culture Producing Nisin Z and Exopolysaccharides in a Fresh Cheese Model,” International Dairy Journal, Vol. 17, No. 2, 2007, pp. 123132.

[24]   V. Monnet, D. Atlan, C. Beal, M. C. Champomier-Verges, M. P. Chapot-Chartier, H. Chouayekh, M. Cogain-Bousquet, M. Deghorain, P. Gaudu, C. Gilbert, E. Guedon, I. Guillouard, P. Goffin, J. Guzzu, P. Hols, V. Juillard, V. Ladero, N. Lindley, S. Lortal, P. Loubiere, E. Maguin, C. Monnet, F. Rul, R. Tourdot-Marechal and M. Yvon, “Métabolisme et Ingénierie Métabolique,” In: G. Corrieu and F. M. Luquet, Eds., Bactéries Lactiques—De la Génétique aux Ferments, Tec & Doc Editions, Paris, 2008, pp. 271-509.

[25]   E. Franciosi, L. Settanni, A. Cavazza and E. Poznanski, “Biodiversity and Technological Potential of Wild Lactic Acid Bacteria from Raw Cows’ Milk,” International Dairy Journal, Vol. 19, No. 1, 2009, pp. 3-11. doi:10.1016/j.idairyj.2008.07.008

[26]   N. Desmasures and E. Beuvier, “Ce qu’il Faut Savoir Avant d’Intervenir sur les Microflores des laits,” In: Microflore du lait cru, CNAOL and GIS Alpes Jura, 2011, pp. 13-74.

[27]   C. Jans, C. Lacroix and L. Meile, “A Novel Multiplex PCR/RFLP Assay for the Identification of Streptococcus bovis/Streptococcus Equinus Complex Members from Dairy Microbial Communities Based on the 16S rRNA Gene,” FEMS Microbiology Letters, Vol. 326, No. 2, 2012, pp. 144-150. ,

[28]   A. Mallet, M. Guéguen, F. Kauffmann, C. Chesneau, A. Sesboué and N. Desmasures “Quantitative and Qualitative Microbial Analysis of Raw Milk Reveals Substantial Diversity Influenced by Herd Management Practices,” International Dairy Journal, Vol. 27, No. 1-2, 2012, pp. 13-21. ,

[29]   A. Badis, D. Guetarni, B. Moussa Boudjemac, D. E. Hennic and M. Kihalc, “Identification and Technological Properties of Lactic Acid Bacteria Isolated from Raw Goat Milk of Four Algerian Races,” Food Microbiology, Vol. 21, No. 5, 2004, pp. 579-588. doi:10.1016/