3.2.3. Tannins

Some red grape cultivars, such as Merlot, Tannat, and Zinfandel, may experience grea<sup>t</sup> difficulty undergoing successful MLF [47,48]. This may be related to certain grape tannins exerting a negative influence on the growth and survival of wine LAB, and consequently on the MLF. Research has been conducted exploring the impact of polyphenols on the growth and viability of wine LAB and their ability to degrade l-malic acid, often with inconsistent results. Polyphenols can have either stimulatory or inhibitory effects on the growth of wine LAB, depending on their type and concentration, and on the selected wine LAB strain in question. Figueiredo et al. [49], Chasseriaud et al. [50], and Stivala et al. [51] showed that tannin compositions containing a high percentage of condensed tannins can strongly affect the viability of *O. oeni* cells, whereas tannin blends consisting of anthocyanins and condensed tannins or catechin and epicatechin monomers and dimers, respectively, only slowed down the growth of bacteria when they were used at the highest concentration. These results are also in agreemen<sup>t</sup> with previous studies that showed no effect or a stimulatory effect of these compounds [49,52,53]. The successive activity of a 3-O-galloyl esterase and gallate decarboxylase, as it has been found in *L. plantarum* [54], may explain the stimulation by the addition of grape tannins.

## 3.2.4. Nutrient Deficiencies

In order to successfully complete MLF, proper nutrition for the wine LAB is of the utmost importance, because wine LAB are characterized as having complex nutritional requirements. Contrary to the fermentation yeas<sup>t</sup> *Saccharomyces cerevisiae*, the bacteria *O. oeni* and other wine LAB cannot utilize inorganic nitrogen sources. Instead, sufficient amounts of organic nitrogen in the form of amino acids and peptides must be supplied. The vitamins pantothenic acid, thiamine, and biotin, as well as the trace elements Mg, Mn, and K, must also be provided to ensure bacterial growth and malolactic activity. Terrade and Orduña [55] investigated the essential growth requirements of four strains of wine LAB from the genera *Oenococcus* and *Lactobacillus*. The two *Oenococcus oeni* strains revealed a larger number of auxotrophies (18 and 23), the two *Lactobacillus* strains only had 11 and 14 auxotrophies. Despite the complex nutritional needs of wine LAB, the amounts they require are, in fact, quite small. Normally, the amount of nutrients contained in the wine matrix is sufficient to meet the needs of LAB, but certain vinification practices can result in nutrient deficiency.

#### 3.2.5. Residual Lysozyme Activity and Chitosan Formulation

If lysozyme is used to control indigenous LAB during the production of wine, it is possible that residual levels of this enzyme may impact the duration of the subsequent MLF [56]. In most cases, racking the wine off the gross lees is recommended. Strains of *O. oeni* are more sensitive to the effects of lysozyme than strains of *Lactobacillus* or *Pediococcus* are.

Certain forms of chitosan, a natural polymer derived from chitin, exhibit antimicrobial properties. Chitosan is well known for its antimicrobial properties against yeast, bacteria, and fungi [57,58], and a preparation extracted from a fungal source has been used to neutralize contamination by *Brettanomyces bruxellensis* [59]. More recently, Chitosanglucan formulation has been released to inhibit wine lactic acid bacteria and delay or inhibit malolactic fermentation. To ensure timely MLF, as well as wine protection, the application of chitosan is recommended at the completion of MLF.
