Enterococcus lactis sp. nov., from Italian raw milk cheeses

Abstract

Ten atypical Enterococcus strains were isolated from Italian raw milk cheeses. The 16S rRNA gene, phenylalanyl-tRNA synthase alpha subunit (pheS), RNA polymerase alpha subunit (rpoA) and the 16S-23S rRNA intergenic transcribed spacer (ITS) sequences, randomly amplified polymorphic DNA (RAPD) PCR and the phenotypic properties revealed that the isolates represent a novel enterococcal species. On the basis of 16S rRNA gene sequence analysis, the isolates were closely related to Enterococcus hirae ATCC 8043 T , Enterococcus durans CECT 411 T andEnterococcus faecium ATCC 19434 T , with 98.8, 98.9 and 99.4 % sequence similarity, respectively. On the basis of sequence analysis of the housekeeping gene pheS, the reference strain, BT159T , occupied a position separate from E. faecium LMG 16198. The group of isolates could be easily differentiated from recognized species of the genus Enterococcus by 16S-23S rRNA ITS analysis, RAPD-PCR and phenotypic characteristics. The name Enterococcus lactis sp. nov. is proposed, with BT159 T (5DSM 23655 T 5LMG 25958 T ) as the type strain.Enterococci belong to a group of organisms known as lactic acid bacteria and large numbers are usually present in vegetables, plant material and foodstuffs, especially those of animal origin such as dairy products (Giraffa, 2003;Fisher & Phillips, 2009). Studies on the microbiota of traditional cheeses of Mediterranean countries, produced mainly from raw milk from sheep, goats or cows, indicate that enterococci are a relevant component of the natural cultures involved in fermentation and that they play an important role in cheese ripening, hence contributing to the cheeses' typical taste and flavour (Foulquié Moreno et al., 2006). In addition, enterococci are also used to extend the shelf life and improve the hygienic safety of foodstuffs because they produce antimicrobial substances such as lactic acid, hydrogen peroxide and bacteriocins (enterocins). Bacteriocins have become of great interest as they are frequently active against several Gram-positive, food-borne pathogens such as Listeria monocytogenes, Staphylococcus aureus and Clostridium botulinum and it has been suggested that they should be used as biopreservatives in foods (Franz et al., 2007). Enterococci have also been investigated for their potential as probiotics, a role that is still controversial as some strains isolated from foods and human samples have been associated with human infections, several virulence factors and antibiotic resistance including resistance to glycopeptide antibiotics (vancomycin) (Foulquié Moreno et al., 2006).Since the revival of the genus Enterococcus by Schleifer & Kilpper-Bälz (1984), phylogenetic studies based on 16S rRNA gene sequences have established the presence of some species groups, namely 'faecium', 'faecalis', 'avium', 'gallinarum' and a fifth group comprising Enterococcus columbae and Enterococcus cecorum. Other described enterococci form individual lines of descent (Devriese et al., 1993;Devriese & Pot, 1995;Stiles & Holzapfel, 1997;Fortina et ...

PreliminaryIn VitroEvaluation of the Probiotic Potential of the Bacteriocinogenic StrainEnterococcus lactisPMD74 Isolated from Ezine Cheese

“…Among these colonies, PMD74 was selected for further experiments as it exhibited the highest antimicrobial activity. e characteristic properties (Gram-positive, catalase-negative and aesculin-positive, coccus-shaped isolate that was able to grow effectively at 10-45°C, pH 9.6, and in the presence of both NaCl and bile salts) of the strain were consistent with the general characteristics of genus Enterococcus [17]. According to the results of the API 20 Strep test, the isolate was observed to generate a positive reaction for Voges-Proskauer, pyrrolidonyl arylamidase, and leucine aminopeptidase, while it was negative for α-galactosidase, β-glucuronidase, and hippurate.…”

“…e strain possessed weak acid production ability from starch. e finding that the PMD74 strain did not produce gas from sucrose, while it produced gas from salicin, also supported the proposition that PMD74 was a different strain than E. hirae [17]. According to the complete 16S rRNA gene sequence analysis, which was performed in order to ensure molecular identification, the isolate was designated as an Enterococcus lactis strain.…”

“…In this respect, the research works on the isolation of novel enterococcal strains other than E. faecalis as well as on their biotechnological and safety potentials hold great importance, especially in the Mediterranean nations. E. lactis, which was first isolated in Russian sour milk products [15], and later in South African fresh sheep milk [16], was described by Morandi et al [17] as a novel species belonging to genus Enterococcus. In several studies conducted by different research groups, the biotechnological, probiotic, and safety-related properties, as well as the bacteriocin-producing potential of E. lactis, have been investigated [6,8,9,18,19].…”

The genus Enterococcus as probiotic: safety concerns

“…The genus Enterococcus has been the target of several studies for use as probiotics (Sivieri et al 2008;Kuritza et al 2011). They have some desirable characteristics for this purpose such as resistance to gastric juice and bile salts (Rossi et al 2003) and production of antimicrobial compounds such as enterocin (Franz et al 1999;Saarela et al 2000).The genus has been isolated from different niches such as water (Oliveira and Pinhata 2008), plants (Svec et al 2011), animals (Jung et al 2007) and foods (Gomes et al 2008) and different species have been used as starter or adjunct culture in the manufacture of fermented products such as cheese (Centeno et al 1999;Ghairi et al 2008;Williams and Withers 2010;Morandi et al 2011) and sausages (Sabia et al 2002;Tanasupawat et al 2008), among others.Although some studies have corroborated its effectiveness as probiotic (Bellomo et al 1980;Saavedra et al 2001;Benyacoub et al 2003;Rivera-Espinoza and Gallardo-Navarro 2010), others have shown correlation of the genus with diseases, which include urinary tract infections (Protonotariou et al 2010), bacteremia (Shankar et al 1999, Shankar et al 2002Tan et al 2010) and endocarditis (Conde-Estéves et al 2010;Rasmussem et al 2010;Heikens et al 2011). However, the host is usually patients with serious diseases and immuno-compromised (Brilliantova et al 2010) what emphasizes its opportunist nature.…”

Isolation, identification, and technological characterization of wild leuconostocs and lactococci for traditional Raib type milk fermentation

“…may belong to species Enterococcus lactis, as these isolates were genotypically related to E. faecium. This species, recently proposed for enterococci isolated from different dairy products (Bauer et al 2009;Morandi et al 2011a;Sukhodolets et al 2005), has been recently validated (Morandi et al 2011b).…”