La dimensión del agua como derecho fundamental

Virulence factors are mainly detected among clinical enterococcal isolates, although studies done on the prevalence of virulence traits among enterococcal strains isolated from food suggest that some strains harbour virulence traits as well [62]. In this regard, in 2001 Eaton and Gasson [50] showed through PCR and gene screening tests, that enterococcal virulence factors were present in clinical, food and starter culture isolates. Though, the prevalence was higher among clinical strains, followed by food isolates; the lowest prevalence was observed for starter isolates [50].

Among enterococcal species, according to Franz et al. [61] and Eaton and Gasson [50], E. faecalis generally harbour more and multiple virulence determinants and with much higher frequencies than E. faecium. Eaton and Gasson found that all of the clinical, food and starter E. faecalis isolates they tested possessed multiple determinants (between 6 and 11), while E. faecium isolates were generally free of virulence determinants, with notable exceptions [50]. In turn, Franz et al. found that of the 47 E. faecalis isolates of food origin they have tested, 78.7% were positive for one or more virulence determinants, compared to 10.4% of the 48 E. faecium isolates of food origin tested [61]. On the other hand, in the Franz et al. study, the isolates exhibiting virulence traits were not necessarily positive for all traits; thus, the prevalence of virulence factors may be considered to be strain or isolate specific [61]. In a similar manner, the Eaton and Gasson results showed that their identified virulence determinants had not previously

been identified, and that this may have resulted from regional differences, suggesting a strain or isolate specificity as well [50].

In 2003, Mannu et al. [124] conducted a study where virulence traits among 94 E. faecium isolates were searched. For the study, 40 isolates were obtained from 3 different traditional goat’s and ewe’s raw milk cheeses produced in the island of Sardinia, 26 from faeces of Sarda breed sheep, and 28 from different clinical samples from patients from different wards staying at one Sardinian hospital. The results demonstrated that of all the isolates the ones obtained from cheeses harboured less virulence determinants than those obtained from patients samples [124]. It was also found that there was a difference in the type of virulence determinants present in cheese and clinical isolates (EfaAfs was the trait isolated in cheeses while Esp was found in clinical samples). No virulence traits were found in sheep faeces strains. The study also revealed that, although there was a clear difference in the type of virulence factor present in isolates of different origin, each E. faecium isolate did not carry more than one virulence determinant, something that the investigators considered as low in prevalence [124]. The results of this study therefore suggested that E. faecium from traditional Sardinian raw goat’s and ewe’s milk cheese should not be considered as potential ‘virulence carriers’ for humans, since only one virulence determinant was found in each cheese positive isolate, and, overall, they were different to the type of virulence determinant isolated in patients [124].

In respect of what type of virulence traits are frequently found in E. faecalis or E. faecium, most of the findings of the Mannu et al. study [124] agree with the ones obtained in previous studies [38, 50, 61, 174]. For example, in the Mannu et al. study [124], the gene for the gel virulence factor was not found in E. faecium strains; this result that was also obtained in the study carried out by Franz et al. [61], while Eaton and Gasson have only found one clinical E. faecium isolate harbouring this virulence trait [50]. All these findings in turn agree with Coque et al. [38], who actually only found this gene in E. faecalis strains. AS was also not found in the Mannu et al. study [124], which agrees with the fact that previously AS has only been described in E. faecalis isolates as well [50, 61, 174]. With regards to Esp, it is not surprising that the Mannu et al. [124] study had found 21 clinical isolates carrying this virulence trait, since Eaton and Gasson [50] and Franz et al. [61] also found Esp in clinical E. faecium strains only. The EfaAfs found in the 19 cheese isolates in the Mannu et al. study [124]

also agrees with those results of Eaton and Gasson [50], who found that this was the only virulence trait present in food strains of E. faecium.

Therefore, according to Franz et al. [61, 62], Eaton and Gasson [50], and Mannu et al. [124], among enterococcal species, E. faecalis of clinical origin generally harbour more and multiple virulence determinants and with much higher frequencies than E. faecium, which are generally free of them. In E. faecium isolates of food origin, only a few have been recognised as producing either cyl (8.3%), Esp (2.1%) [61], or EfaAfs [124]. E. faecium appears to pose a lower risk for use in foods since their strains are generally free of virulence determinants [50, 61, 62, 124].

Table 8 shows the results of Mannu et al. study [124] of E. faecium strains. Table 9 shows the most common virulence factors found in E. faecalis and E. faecium.

Table 8: E. faecium strains isolated from cheese, sheep, and hospitalised patients, with their PCR results for virulence traits genes (summarised according to [124]).

Product/Origin Number of positive (+) isolates to

virulence determinants Number of

isolates

tested Ace EfaAfs GelE AS Esp

(a) Ewe’s cheeses

Casu Axedua 2 - - - Fiore Sardob 30 - 16 - - - Pecorino Sardoc 8 - 3 - - - Total 40 0 19 0 0 0 (b) Sheep Sheep's faeces 26 - - - Total 26 0 0 0 0 0 (c ) Nosocomial patients Respirator 11 - - - - 10 Drain 5 - - - - 1 Anal tampon 3 - - - - 2 Skin tampon 1 - - - - 1 Pus 1 - - - - 1 Arterial catheter 1 - - - - 1 Vesical catheter 2 - - - - 2 Expectoration 1 - - - Bronchial lavage 1 - - - - 1 Urine 2 - - - - 2 Total 28 0 0 0 0 21

a. Goat's milk fresh cheese

b. Hard uncooked ewe's milk cheese c. Semi-cooked ewe's milk cheese

Table 9: Common enterococcal virulence traits and the enterococcal species where they have been found

Virulence factor Enterococcal

species References

AS E. faecalis Jett et al. [91]

Süßmuth et al. [174] Elsner et al. [53] Franz et al. [61] Eaton and Gasson [50] E. faecium Jett et al. [91]

Elsner et al. [53]

Esp E. faecium Eaton and Gasson [50]

Franz et al. [61]

Mannu et al. [124]

Ace E. faecalis Nallapareddy et al. [140, 141]

Cyl E. faecalis Jett et al. [91]

Huycke et al. [88]

Elsner et al. [53]

E. faecium Jett et al. [91] Franz et al. [61]

EfaAfs E. faecalis Lowe et al. [117]

E. faecium Singh et al. [169] Eaton and Gasson [50]

Mannu et al. [124]

Gel E. faecalis Jett et al. [91]

Kuhnen et al. [105]

Coque et al. [38]

Elsner et al. [53]

Franz et al. [61]

E. faecium Eaton and Gasson13 [50]