Influence of the storage temperature of raw sheep milk on the spoilage potential of Pseudomonas spp.

Abstract

Due to their quality, high yield and nutritional value, sheep dairy products have been gaining market space and attracting new producers. Pseudomonas spp. is the most common psychrotrophic genus present in bovine refrigerated raw milk and is characterized by intense lipoproteolytic activity. Metalloprotease aprX is one of the enzymes produced by Pseudomonas spp. which has great industrial importance, since it has specific activity in the deterioration of casein, resulting in significant changes in physicochemical properties and sensorial characteristics of raw milk and dairy products. The aim of this study was to observe the occurrence of Pseudomonas spp. in raw sheep milk and evaluate the influence of storage temperature on the spoilage potential of the strains. Raw sheep milk samples were stored at 4 °C or 9 °C (72 h) and plated in Pseudomonas Agar Base supplemented with CFC (25 ºC / 48 h). The isolates characteristic of Pseudomonas had their genus, species (P. putida, P. fluorescens and P. aeruginosa), and presence of gene aprX confirmed by Polymerase Chain Reaction (PCR). The lipoproteolytic capacity and the spoilage potential of the isolates were evaluated in Milk Agar (10%) and Tributyrin Agar Base (1%) (21 °C / 72 h). Initial (2 h) populations of Pseudomonas spp. ranged from 1.0 to 4.2 log CFU/mL; after 72 h, the values observed in milk samples stored at 9 °C (3.0–7.0 log CFU/mL) were higher than those obtained in samples at 4 °C (1.6–6.0 log CFU/mL). A total of 255 strains of Pseudomonas spp. were isolated: 30.2% (77) from milk stored at 4 °C and 69.8% (178) from milk at 9 °C. At both temperatures, P. fluorescens was predominant, followed by P. putida. P. aeruginosa strains, isolated only from samples stored at 9 °C, showed high lipoproteolytic capacity and were characterized by higher proteolytic potential than lipolytic. P. fluorescens was the only species to present high proteolytic (halos > 2 cm) potential. Considering a single storage temperature (4 °C and 9 °C), only P. fluorescens showed higher proteolytic potential than lipolytic. The aprX gene was predominant in P. fluorescens and P. putida isolates (4 °C and 9 °C); strains obtained from milk stored at 4 °C showed higher proteolytic potential than those isolated from milk at 9 °C. All strains of P. fluorescens and only 46.42% of P. putida presented proteolytic capacity at 9 °C. The aprX gene was observed in all P. aeruginosa strains, but only 95.65% of them showed proteolytic capacity. The lipolytic potentials of P. fluorescens and P. putida were not influenced by milk storage temperature. P. fluorescens was the only species to present high lipolytic potential. P. putida isolated from milk at 9 °C was the only species to present higher lipolytic potential than proteolytic. According to the microbiota and the storage temperature, there may be a predominance of proteolytic or lipolytic enzymes in milk and consequently, defects in sheep dairy products. It should also be considered that most of the isolates of Pseudomonas spp. presented lipoproteolytic capacity, which emphasizes the importance of low initial populations of this microorganism in raw sheep-chilled milk, mainly when used in the production of dairy products.