Editing Malolactic fermentation (section)

==Lactic acid bacteria==
[[File:O. oeni.jpg|right|thumb|''Oenococcus oeni'']]
All lactic acid bacteria (LAB) involved in winemaking, whether as a positive contributor or as a source for potential faults, have the ability to produce lactic acid through the metabolism of a sugar source, as well as the metabolism of L-malic acid. Species differ in how they metabolise the available [[sugars in wine]] (both [[glucose]] and [[fructose]], as well as the unfermentable [[pentose]]s that [[wine yeast]]s do not consume). Some bacteria species use the sugars through a [[homofermentative]] pathway, meaning only one main [[Product (biology)|end product]] (usually lactate) is produced, while others use [[heterofermentative]] pathways that can create multiple end products such as carbon dioxide, [[ethanol]], and [[acetate]]. While only the L-[[isomer]] of lactate is produced by LAB in the conversion of malic acid, both hetero- and homofermenters can produce D-, L- and DL-isomers of lactic from glucose which may contribute to slightly different sensory properties in the wine.<ref name="Jacobson"/>

While ''O. oeni'' is often the LAB most desired by winemakers to complete malolactic fermentation, the process is most often carried out by a variety of LAB species that dominate the must at different points during fermentations. Several factors influence which species will be dominant, including fermentation temperature, nutritional resources, the presence of [[sulfur dioxide]], interaction with yeast and other bacteria, pH, and alcohol levels (''Lactobacillus'' species, for example, tend to prefer higher pH and can tolerate higher alcohol levels than ''O. oeni''), as well as initial inoculation (such as "wild" ferments versus an inoculation of cultured ''O. oeni'').<ref name="Yair"/>

===''Oenococcus''===
[[File:Malolactic inoculation and nutrient.JPG|left|thumb|Cultured ''Oenococcus oeni'' inoculum strain and "Opti-malo" nutrient additive]]
The genus ''[[Oenococcus]]'' has one main member involved in winemaking, ''O. oeni'', once known as ''Leuconostoc oeni''. Despite having the name ''Oeno[[coccus]]'', under the microscope, the bacterium has a [[bacillus (shape)]] rod shape. The bacteria is a [[Gram-positive]], [[facultative anaerobe]] that can utilize some [[oxygen]] for [[aerobic respiration]] but usually produces cellular energy through fermentation. ''O. oeni'' is a heterofermenter that creates multiple end products from the use of glucose with D-lactic acid and carbon dioxide being produced in roughly equal amounts to either ethanol or acetate. In [[reductive condition]]s (such as near the end of alcoholic fermentation), the third end product is usually ethanol while in slightly [[oxidative]] (such as early in alcohol fermentation or in an [[ullage (wine)|untopped barrel]]), the bacteria are more likely to produce acetate.<ref name="Boulton"/>

Some ''O. oeni'' strains can use fructose to create [[mannitol]] (which can lead to wine fault known as mannitol taint), while many other strains can break down the [[amino acid]] [[arginine]] (which can be present in the wine that is resting on the [[lees (wine)|lees]] after fermentation from the [[Autolysis (wine)|autolysis]] of dead yeast cells) into [[ammonia]].<ref name="Wine Micro"/>

In addition to the [[hexose]] glucose and fructose sugars, most strains of ''O. oeni'' can use the [[residual sugar|residual pentose]] sugars left behind from yeast fermentation including [[L-arabinose]] and [[ribose]]. Only around 45% of ''O. oeni'' strains can ferment [[sucrose]] (the form of sugar usually added for [[chaptalization]] that gets converted by yeast into glucose and fructose).<ref name="Wine Micro"/>

Winemakers tend to prefer ''O. oeni'' for several reasons. First, the species is compatible with the main wine yeast ''[[Saccharomyces cerevisiae]]'', though in cases where both MLF and alcoholic fermentation are started together, the yeast most often outcompetes the bacterium for nutritional resources which may cause a delay in the onset of malolactic fermentation. Second, most strains of ''O. oeni'' are tolerant to the low pH levels of wine and can usually deal with the standard [[alcohol level]]s that most wines reach by the end of fermentation. Additionally, while sulfur dioxide levels above 0.8 molecular SO<sub>2</sub> (pH dependent but roughly 35-50 ppm) will inhibit the bacteria, ''O. oeni'' is relatively resistant compared to other LAB. Finally, ''O. oeni'' tends to produce the least amount of [[biogenic amines]] (and most lactic acid<ref name="Jacobson"/>) among the lactic acid bacteria encountered in winemaking.<ref name="Boulton"/>

===''Lactobacillus''===
[[File:20101210 014809 LactobacillusBulgaricus.jpg|right|thumb|''Lactobacillus'' from a yogurt sample.]]
Within the genus ''Lactobacillus'' are both heterofermentative and homofermentative species. All lactobacilli involved in winemaking are Gram-positive and [[microaerophilic]], with most species lacking the enzyme [[catalase]] needed to protect themselves from [[oxidative stress]].<ref name="Wine Micro"/>

Species of ''Lactobacillus'' that have been isolated from wine and grape must samples across the globe include ''[[L. brevis]], [[L. buchneri]], [[L. casei]], [[L. curvatus]]'', [[L. delbrueckii subsp. lactis|''L. delbrueckii'' subsp. ''lactis'']], ''[[L. diolivorans]], [[L. fermentum]], [[L. fructivorans]], [[L. hilgardii]], [[L. jensenii]], [[L. kunkeei]], [[L. leichmannii]], [[L. nagelii]], [[L. paracasei]], [[Lactobacillus plantarum|L. plantarum]]'', and ''[[L. yamanashiensis]]''.<ref name="Wine Micro"/>

Most ''Lactobacillus'' species are undesirable in winemaking with the potential of producing high levels of [[volatile acidity]], off odors, [[wine haze]], gassiness, and [[sediment (wine)|sediment]] that can be deposited in the bottle, especially if the wine had not been [[filtered (wine)|filtered]]. These bacteria also have the potential to create excessive amounts of lactic acid which can further influence the flavor and sensory perception of the wine. Some species, such as the so-called "ferocious ''Lactobacillus''", have been implicated in causing sluggish or [[stuck fermentation]]s, while other species, such as ''L. fructivorans'', have been known to create a cottony [[mycelium]]-like growth on the surface of wines, nicknamed "Fresno mold" after the wine region where it was discovered.<ref name="Boulton"/>

===''Pediococcus''===
[[File:Acrolein.svg|left|thumb|Acrolein taint is a common wine fault that undesirable species of ''Pediococcus'' can introduce to wine. Acrolein can interact with various phenolic compounds, imparting a bitter taste to the wine.]]
So far, four species from the genus ''Pediococcus'' have been isolated in wines and grape must, ''[[P. inopinatus]], [[P. pentosaceus]], [[P. parvulus]]'', and ''[[P. damnosus]]'', with the last two being the species most commonly found in wine. All ''Pediococcus'' species are Gram-positive with some species being micro-aerophilic while others utilizing mostly aerobic respiration. Under the microscope, ''Pediococcus'' often appear in pairs of pairs or tetrads which can make them identifiable. Pediococci are homofermenters, metabolizing glucose into a [[racemic mixture]] of both L- and D-lactate by [[glycolysis]].<ref name="Zoecklein"/> However, in the absence of glucose, some species, such as ''P. pentosaceus'', begin using [[glycerol]], degrading it into [[pyruvate]] which later can be converted to diacetyl, acetate, [[2,3-butanediol]] and other compounds that can impart unfavorable characteristics to the wine.<ref name="Wine Micro"/>

Most ''Pediococcus'' species are undesirable in winemaking due to the high levels of diacetyl that can be produced, as well as increased production of biogenic amines that has been implicated as one potential cause for [[red wine headache]]s. Many species of ''Pediococcus'' also have the potential to introduce off odors or other wine faults to the wine such as the bitter-tasting "acrolein taint" that comes from degradation of glycerol into [[acrolein]] which then reacts with [[phenolic compounds in the wine]] to produce a bitter-tasting compound.<ref name="Boulton"/>

One species, ''P. parvulus'', has been found in wines that have not gone through MLF (meaning malic acid is still present in the wine), but has still had its bouquet altered in a way that enologist have described as "not spoiled" or flaw. Other studies have isolated  ''P. parvulus'' from wines that have gone through malolactic fermentation without the development of off odors or wine faults.<ref name="Wine Micro"/>

===Nutritional requirements===
[[File:Sediment at bottom of wine barrel.jpg|right|thumb|When yeast cells die, they sink to the bottom of the tank or barrel, creating the "lees" sediment visible in this picture. The autolysis of the dead yeast cells is a source of a nutrients for lactic acid bacteria.]]
Lactic acid bacteria are [[fastidious organism]]s that cannot synthesize on their own all of their complex nutritional requirements. For LAB to grow and complete malolactic fermentation, the constitution of the wine medium must provide for their nutritional needs. Like wine yeast, LAB require a carbon source for energy metabolism (usually sugar and malic acid), nitrogen source (such as amino acids and [[purine]]s) for [[protein synthesis]], and various vitamins (such as [[Niacin (substance)|niacin]], [[riboflavin]], and [[thiamine]]) and minerals to assist in the synthesis of enzymes and other cellular components.<ref name="Zoecklein"/>

The source for these nutrients is often found in the grape must itself, though MLF inoculations that run concurrent with alcoholic fermentation risk the yeast outcompeting the bacteria for these nutrients. Towards the end of fermentation, while most of the original grape must resources have been consumed, the [[lysis]] of dead yeast cells (the "lees") can be a source for some nutrients, particularly amino acids. Plus, even "dry" wines that have been fermented to dryness still have [[residual sugars|unfermentable pentose sugars]] (such as arabinose, ribose and [[xylose]]) left behind that can be used by both positive and spoilage bacteria. As with wine yeast, manufacturers of cultured LAB inoculum usually offer specially prepared nutritional additives that be used as a supplement. However, [[YAN (wine)|unlike wine yeast]], lactic acid bacteria can not use the supplement [[diammonium phosphate]] as a nitrogen source.<ref name="Wine Micro"/>

Before the introduction of complex nutritional supplements and advances in freeze-dried cultures of LAB, winemakers would cultivate their inoculum of lactic acid bacteria from [[Microbiological culture|culture slants]] provided by laboratories. In the 1960s, these winemakers found it easier to create [[starter cultures]] in media that contained [[apple juice|apple]] or [[tomato juice]]. This "tomato juice factor" was discovered to be a derivative of [[pantothenic acid]], an important [[growth factor]] for the bacteria.<ref name="Boulton"/>

As with yeast, oxygen can be considered a nutrient for LAB, but only in very small amount and only for microaerophilic species such as ''O. oeni''. However, no evidence exists currently to suggest that malolactic fermentation runs more smoothly in aerobic conditions than in complete anaerobic conditions, and in fact, excessive amounts of oxygen can retard growth of LAB by favoring conditions of competing microbes (such as ''Acetobacter'').<ref name="Boulton"/>

===Native LAB species in the vineyard and the winery===
[[File:Harvested_grapes_being_loaded_into_crusher_destemmer.jpg|left|thumb|Winery equipment offers multiple contact points for native populations of lactic acid bacteria to be introduced to the wine.]]
''Oenococcus oeni'', the LAB species most often desired by winemakers to carry out malolactic fermentation, can be found in the vineyard, but often at very low levels. While moldy, damaged fruit has the potential to carry a diverse flora of microbes, the LAB most often found on clean, healthy grapes after [[harvest (wine)|harvest]] are species from the ''Lactobacillus'' and ''Pediococcus'' genera. After crushing, microbiologists usually find populations under 10<sup>3</sup> [[colony forming units]]/mL containing a mix of '' P. damnosus, L. casei, L. hilgardii,'' and '' L. plantarum'', as well as ''O. oeni''. For musts that do not receive an early dose of sulfur dioxide to "knock back" these wild populations of LAB, this flora of bacteria compete with each other (and the wine yeasts) for nutrients early in fermentation.<ref name="Boulton"/>

In the winery, multiple contact points can be home to native population of LAB including [[oak barrels]], pumps, hoses, and bottling lines. For wines where malolactic fermentation is undesirable (such as fruity white wines), the lack of proper [[sanitation]] of wine equipment can lead to the development of unwanted MLF and result in [[wine fault]]s. In cases of oak barrels where full and complete sanitation is almost impossible, wineries often mark barrels that have contained wines going through MLF and keep them isolated from "clean" or brand new barrels that they can use for wines that are not destined to go through MLF.<ref name="Yair"/>

===''Schizosaccharomyces'' yeast===
Several species in the genus ''[[Schizosaccharomyces]]'' use L-malic acid, and enologists have been exploring the potential of using this wine yeast for deacidifying wines instead of the traditional route of malolactic fermentation with bacteria. However, early results with ''[[Schizosaccharomyces pombe]]'' have shown a tendency of the yeast to produce off odors and unpleasant sensory characteristics in the wine. In recent years, enologists have been experimenting with a [[mutant]] strain of ''[[Schizosaccharomyces malidevorans]]'' that has so far been shown to produce less potential wine flaws and off odors.<ref name="Wine Micro"/>