Bioactive Compounds from and against Yeasts in the One Health Context: A Comprehensive Review

Abstract

Yeasts are the most used microorganisms for biotechnological purposes. Although they have been mainly recognized for their application in the beverage and bioethanol industries, these microorganisms can be efficiently employed in pharmaceutical and food production companies. In these industrial sectors, yeasts are highly desirable for their capacity to produce bioactive compounds from simple substrates, including wastes. In this review, we present the state of the art of bioactive compound production in microbial cell factories and analyze the avenues to increase the productivity of these molecules, which benefit human and environmental health. The article addresses their vast biological activities, from preventing to treating human diseases and from pre to postharvest control on agroindustrial streams. Furthermore, different yeast species, genetically engineered or not, are herein presented not only as biofactories of the referred to compounds but also as their targets. This comprehensive analysis of the literature points out the significant roles of biodiversity, bioprospection, and genome editing tools on the microbial production of bioactive compounds and reveals the value of these approaches from the one health perspective.

1. Introduction

The concept of one health recognizes that human health relies on the health of other living beings and the environment [1]. In this context, considering the biotechnological potential of yeasts, alternatives to combating bacterial antibiotic resistance, improving the nutritional content of foods, increasing sustainable management of crops, and replacing conventional raw materials with residual biomass are envisaged, thus ensuring a more sustainable production chain. As stated by Elnaiem et al. [2], there is a need for urgent investment in strategies that reduce the risk of the appearance of new diseases, which will improve animal, human and environmental health and act in the fight against climate change.

Plant-associated yeasts can control their host plant’s microbiota, thus presenting as an alternative to synthetic pesticides and as inducers of vegetal growth. They are, therefore, potential substitutes for conventional fertilizers or growth hormones. Additionally, second-generation biorefinery strategies have shown proven efficiency in converting residual substrates (e.g., agroindustrial wastes) into a myriad of bioproducts through the metabolism of yeasts, whether genetically engineered or not [3,4,5,6,7,8,9,10,11,12,13]. Therefore, the bioactive compounds produced by yeasts allow the achievement of the one health goals, as they bring benefits not only to human health but also to animal and plant health, helping to maintain environmental balance. In the following sections, we present a comprehensive review of the most valuable bioactive compounds produced by (and against) yeasts and how they can contribute to a more sustainable future.

2. Bioactive Compounds Naturally Produced by Non-Engineered Yeasts

Polysaccharides, polyphenols, carotenoids, flavonoids, and terpenoids are part of the wide range of compounds that have biological activities, including antimicrobial, immunosuppressive, anticancer, and anti-inflammatory activity [14,15]. Although some of them are complex molecules, these bioactive compounds can be produced by microorganisms through biosynthetic processes from simple substrates [16]. Some yeasts have stood out in this context, namely Saccharomyces cerevisiae [17,18], Komagataella phaffii (formerly Pichia pastoris) [19], Hansenula polymorpha [20], Rhodosporidium diobovatum [21], Aureobasidium pullulans [22], Rhodotorula glutinis [23,24], Saccharomyces bayanus, Candida stellata, Kluyveromyces thermotolerants, and Hanseniaspora uvarum [25,26]. Mostly, yeast fermentation products are employed in the nutritional improvement of food products. However, based on the versatility of their metabolic routes, these microorganisms synthesize bioactive compounds of different molecular classes, such as peptides, polyphenols, and terpenoids [27,28].

Table 1. Bioactive compounds produced by different nongenetically modified yeasts, their commercial interest, and market value.

Compound	Formula	Commercial Interest	Market Value in Millions of USD (Year)	Reference
Glutathione		Antioxidant	316.56 (2020)	[17,18]
Melatonin		Sleep cycle receptor mediator and antioxidant	437.9 (2021)	[26,29]
Riboflavin		Food coloring and supplement	397 (2023)	[30,31]
Toxin K1	KEX1 e KEX2 proteases encoded	Antifungal	-	[32]
Toxin K28		Antifungal	-	[33]
Toxin K2		Antifungal	-	[34]
Toxin KP	Decapeptide KP (AKVTMTCSAS)	Toxoplasmosis and antifungal treatment	-	[35]
Rhodotorulic Acid		Iron bioavailability and regulator of iron-mediated membrane transporters	-	[36]
Ferrichrome		Bioavailability and regulator of iron and other metals	-	[36,37]
Acetic Acid		Preservative and food additive; drug production, industrial and laboratory solvent	2900 (2021)	[38,39]
Citronellol		Antifungal	~146.8 (2030)	[40,41]
Phenylethyl Alcohol		Antimicrobial, antiseptic, disinfectant, aromatic essence, and preservative of pharmaceuticals and cosmetics	255.3 (2021)	[40,42]
Indol		Fragrance component and flavor aggregator	35.7 (2021)	[25,43]
Quercetin-3-glucoside		Antiviral, antioxidant	-	[44]
Caffeic Acid		Antioxidant, anti-inflammatory		[44]
Naringenin		Antioxidant, anti-inflammatory, antifungal		[44]

summarizes the main bioactive compounds produced by nongenetically modified yeasts of different species and points out their commercial interests and market values.

2.1. Pharmacological Outlooks

Notably, the main species used in the production of bioactive compounds is Saccharomyces cerevisiae [45,46]. The glutathione tripeptide, formed by a thiol of cysteine, glycine, and glutamine, is a major example of these products [47,48]. Although humans naturally produce it, the significant antioxidant effect of this molecule makes it a product of commercial interest, being sold mainly in retail chains and pharmacies as a supplement [49,50]. The interest in glutathione, however, is not restricted to combating free radicals. In humans, this tripeptide acts in the metabolism of peroxides [51] and xenobiotics [52], the inflammatory/immunological response [53], and the activation of some enzymes [54]. Currently, like most bioactive compounds, glutathione is also mainly produced by strains of S. cerevisiae due to its safety in food production [17,18]. In addition to S. cerevisiae, though, other yeasts can be used for the same purpose, such as the methylotrophic yeasts K. phaffii [19] and H. polymorpha [20], as well as the marine yeast Rhodosporidium diobovatum [21].

Melatonin is another major antioxidant that can be produced by yeast species, such as S. cerevisiae, S. bayanus, and Saccharomyces uvarum. This compound, an indole amine, is part of the composition of wines and beers, being produced naturally during the fermentation process [26,55,56,57,58,59]. Another important role of melatonin in the human and animal body is the maintenance of sleep rhythms [60,61]. Cardinali [62] reports the prevention of neurodegenerative diseases such as Alzheimer’s and Parkinson’s in experimental models when melatonin is administered. Reiter et al. [63] report the reduction of stroke-provoked neural damage in experimental models. Commercially, melatonin is sold as a sleep hormone, being found in drugstores and retail. Despite being naturally produced by the body, it can often be out of balance due to biological factors or habits that inhibit its production—that is why it is commercially sought after as a replacement.

Biologically active molecules can also be peptides that act as toxins depending on the environmental conditions. Bioactive peptides are protein segments abundantly found in yeast extracts, especially those generated by S. cerevisiae, Kluyveromyces marxianus, and Candida utilis, since the cells of these microorganisms harbor 35 to 60% of protein in their dry mass [64]. These yeasts are able to produce antimicrobial peptides that fight other yeasts, bacteria, and molds; in addition, they produce mycocins, extracellular proteins that hinder the growth of Gram-negative and Gram-positive bacteria [65].

The so-called killer yeasts are also extensively studied for the production of bioactive compounds. In addition to acting in the biological control of phytopathogens (as discussed in the following section), these yeasts can serve as producers of new human and animal health drugs. In this context, a killer decapeptide produced by Pichia anomala stands out, showing in vitro and in vivo inhibitory activities against influenza A and human immunodeficiency type-1 (HIV-1) viruses [66,67]. Decapeptides such as this one have already shown activity against infections caused by Candida albicans (also a yeast), thanks to the destruction of its biofilm in invaded tissues [68], and Toxoplasma gondii (the etiological agent of toxoplasmosis) [69].

Tested for pharmacological purposes, the yeast Pichia kudriavzevii showed the production of a toxin with activity against human-infective bacteria, such as Escherichia coli, Enterococcus faecalis, Klebsiella spp., Staphylococcus aureus, Pseudomonas aeruginosa, and Pseudomonas alcaligenes. The toxin produced by P. kudriavzevii has a high specificity of action and high toxicity against the above pathogens, which are extremely desirable characteristics for drug development [70]. Alternatively, the toxin can also be used as a prophylactic measure, preventing these pathogens from reaching humans through the ingestion of contaminated food [71].

Yeasts are also able to accumulate lipids for the pharmaceutical industry [72,73]. The so-called single-cell oils (SCO) include long-chain fatty acids of nutritional interest, such as the polyunsaturated acids Eicosapentaenoic (produced by Meyerozyma guilliermondii), Linoleic (produced by Apiotrichum brassicae, Candida tropicalis, Metschnikowia pulcherrima, P. kudriavzevii, M. guilliermondii, Lodderomyces elongisporus, and Rhodotorula mucilaginosa), and γ-Linolenic (produced by A. brassicae, M. pulcherrima, M. guilliermondii, L. elongisporus, and R. mucilaginosa) [72,74,75,76].

The tropical yeast Saccharomyces boulardii, isolated from lychee and mangosteen peels, has interesting properties for supplementation in the human microbiota, given its ability to adapt to the inhospitable conditions of the gastrointestinal system. This yeast can produce high levels of acetic acid, inhibiting virulent strains of E. coli [38], which are eventually responsible for human food poisoning. Saccharomyces boulardii is also considered efficient in treating inflammatory bowel diseases [77,78] and improving inflammation caused by Citrobacter rodentium [79]. This yeast also demonstrated protection against infection generated by the yeast C. albicans [80].

Riboflavin is also an important bioactive compound produced by yeasts. Capable of preventing deficiency symptoms such as dermatitis, this vitamin also plays a key role in the food industry. Its chemical synthesis requires organic solvents, generates waste that is harmful to the environment, and requires a lot of energy compared to the yeast fermentation process [30]. Currently, the yeast M. guilliermondii stands out in the large-scale production of riboflavin [81]. In addition, the marine yeast Candida membranifaciens subsp. flavinogenie W14-3, isolated from sea water in China, also showed a high potential for producing this bioactive compound [30].

2.2. Microbial Competition and Ecological Food Production

Due to multiple adverse situations, microorganisms are not always inserted in a natural habitat with abundant nutrients from which they can benefit. In these cases, microbial competition is inevitable, and the competition tools of each microorganism can be exploited as agroecological strategies. In this context, yeasts seem to have a prominent place [82,83]. Due to interactions with other microorganisms in natural environments, yeasts can use competitive approaches that guarantee advantages against other opposing organisms, either through mechanisms of enzyme secretion, toxin production, or the release of volatile organic compounds (VOCs) [84].

Volatiles produced by yeasts can display antifungal and bacteriostatic action. Two of these compounds are ethanol and 2,3-butanediol, both produced by the yeasts S. cerevisiae and P. kudriavzevii, which have already been proven capable of reducing the growth of the filamentous fungi Aspergillus flavus and Aspergillus parasiticus [85]. The volatile compound 2-phenylethanol, produced by P. anomala, has also demonstrated a fungus-control effect; in one case, this effect was against Aspergillus ochraceus [86], a well-known generator of toxins in food [87].

The species Sporidiobolus pararoseus has the ability to control the plant illness caused by the gray fungus Botrytis cinerea. Huang et al. [88] attributed this activity to 2-ethyl-1-hexanol. The yeast Candida intermedia was also able to control this phytopathogen by producing the VOCs 1,3,5,7-cyclooctatetraene, 3-methyl-1-butanol, 2-nonanone, and phenylethyl alcohol [89]. Cyberlindnera jadinii, Candida fryrichii, C. intermedia, and Lachancea thermotolerans have also been identified as producers of 2-phenyl ethanol, inhibiting the molds Aspergillus carbonarius and A. ochraceus [90,91,92].

Against postharvest bacterial diseases, the polymorphic yeast Aureobasidium pullulans has been shown to produce secondary metabolites such as aureobasidins, liamocins, 2-propyl acrylic acid, and 2-methylene succinic acid, which act against bacteria such as Salmonella typhi, Proteus vulgaris, E. coli, P. aeruginosa, Klebsiella pneumoniae, Bacillus subtilis, S. aureus, and Sarcina ventriculi. Liamocins also showed selective activity against bacteria of the genus Streptococcus [93,94,95,96]. The biocontrol strategy using VOCs produced by yeasts is interesting to use to maintain the shelf life of fruits such as grapes, especially with compounds produced by Wickerhamomyces anomalus, M. pulcherrima, and A. pullulans [97]. Furthermore, VOCs produced by Candida saque were also efficient in controlling the proliferation of fungi in apples [40].

Marine yeasts are often found in hostile conditions to maintain their lives. Marine species, such as the aforementioned A. pullulans, can produce siderophores—secondary metabolites used by microorganisms as chelators of essential trace elements such as iron [98,99,100,101,102,103]. The so-referred yeast produces a high concentration of the siderophore hydroxamate, a bioactive compound capable of inhibiting the cell growth of the bacteria Vibrio anguillarum and Vibrio parahaemolyticus, which are found in sick marine animals. In fact, siderophores are effective in controlling pathogenic bacteria in marine fish, presenting as an important alternative for pisciculture and the remediation of sites with radioactive waste [104].

In addition to marine yeasts, yeasts associated with plants and fruits can also produce siderophores, such as R. glutinis, Rhodotorula rubra, and species of the genus Meyerozyma, such as M. guilliermondii [23,105]. Although some siderophores have shown pharmacological potential, allowing the control of pathogenic microorganisms [98,101], it is in agriculture that they primarily receive attention. These compounds are quite efficient in protecting plants of agronomic interest against fungal and bacterial infections [99]. The chelating action of these compounds also contributes to the availability of essential micronutrients for plant development [106].

Competition for micronutrients, especially iron, limits the development of microorganisms [83]. The species R. glutinis, isolated from a rhizosphere environment, uses the siderophore rhodotorulic acid to inhibit the growth of its competitor B. cinerea [107]. Along the same line, M. pulcherrima, also found in rhizospheres [108], inhibits its competitors through pulcherriminic acid [109]. Still in the context of the rhizosphere, it should be noted that the growth of plant roots can benefit from the presence of yeasts in the rhizosphere [110,111].

In terms of microbial competition, the production of killer toxins is another yeast strategy. This trait has been widely identified in different yeast genera [112]. In nature, these killer microorganisms are found in different environments, whether in fruits, water, or soil [113]. Toxins can be active against filamentous fungi [114] and bacteria [115]. In shrimp culture, for example, wild yeasts can also confer advantages to the production system. The psychrotolerant yeast Mrakia frigida, found in marine sediments in Antarctica, produces a killer toxin against the shrimp-pathogenic yeast Metschnikowia bicuspidata, and can be inoculated in shrimp breeding tanks to improve this animal’s health [116]. Still in the context of animal production, the yeast Williopsis mrakii deserves attention in silage conservation. While lactic fermentation (by lactic acid bacteria) of silage is desirable, subsequent aerobic consumption of lactic acid (by other microorganisms) causes its deterioration. In this scenario, Lowes et al. [117] demonstrated that the HMK mycocin produced by W. mrakii was able to prevent the development of spoilage microorganisms.

Enzymes secreted by yeasts are also part of the list of mechanisms used to control phytopathogenic fungi. Their antifungal action is often due to their hydrolytic capacity; this is the case, for example, of the chitinases produced by the genera Pichia, Debaryomyces, Metschnikowia, Meyerozyma, and Saccharomycopsis, which act on fungal cell walls composed of chitin. Among the hydrolases, proteases from Aureobasidium, Pichia, and Saccharomycopsis, and glucanases from Wickerhamomyces and Pichia also have a fungicidal or fungistatic effect [118,119,120,121,122].

Strains of W. anomalus and M. guilliermondii have significant effects in reducing the postharvest disease caused by the fungus Colletotrichum gloeosporioides in papaya [123]. The yeast W. anomalus can also control the cherry tomato gray mold caused by the fungus Lycopersicon esculentum [124]. Likewise, R. glutinis strains ensure the significant control of blue rot caused by Penicillium expansum in postharvested apples [125].

Therefore, alternatives for improving the ecological practices of food production arise from the ecological relationships of yeasts. Bioactive compounds produced by yeasts can act in pre and postharvest control, replacing synthetic agricultural defensives, which have a high environmental impact on and pose high environmental risk to human and animal health [113,124].

3. Other Microorganisms’ Bioactive Compounds Acting against Yeast

In natural environments, the production of organic acids such as acetic, lactic, and propionic acids affects the interspecific relationships among microbial cells. The elucidation of their mechanisms of inhibition, though, remains not fully understood. Acetic acid, for example, exhibits a synergistic effect with lactic acid with the capability of preventing fungal/yeast growth, being probably more potent due to its higher pKa value (4.76), which facilitates its uptake by cells and leads to a higher level of acid dissociation in the cytoplasm, damaging the cell [126]. The same holds true for other organic acids—e.g., propionic acid displays a pKa value of 4.87. Mixtures of acids usually appear in the natural ecosystem and have been applied as fungal/yeast inhibitors. A mix of formic, acetic, propionic, butyric, caproic, and n-valeric acid, for example, is naturally produced by lactic acid bacteria (LAB) [127]. On the other hand, in case these acids are thought to be used as biocontrol agents of yeasts, it should be noted that their exacerbated use may lead to a contamination process in soil, plants, water bodies, and air, thus also affecting animal and human health. Thus, there is a demand for new biocompounds in aiming to control yeasts without environmental contamination [128].

In this sense, LAB may also offer alternatives. Reuterin, for example, produced by Lactobacillus reuteri, is a low-molecular-weight compound that exhibits antimicrobial activity against yeasts such as C. albicans [129]. The cyclic dipeptides produced by LAB are also good examples of bioactive compounds that act against yeasts. Lactobacillus plantarum has been cultivated, seeking metabolites to biocontrol mold and yeasts, and two compounds were isolated with this aim: cyclo(L-Phe-L-Pro) and cyclo(L-Phe-trans-4-OH-L-Pro). The former’s minimal inhibitory concentration (MIC) was 20 mg/mL in assays with Penicillium roqueforti and Aspergillus fumigatus. Additionally, a synergistic effect was observed when this compound was associated with phenyllactic acid, which reduced the MIC to 10 mg/mL [130]. Several lactobacilli can produce these compounds, but the mechanism of action and biochemical pathways of cyclic dipeptide acting as inhibitors of fungi are still unknown [131].

Fatty acids (FAs) are also good examples of molecules with antifungal activity. The literature demonstrates that the chain length of the fatty acid appears to be relevant in this role. The FA identified as lauric (C12) and capric (C10) seems to be the most toxic against C. albicans [132]. Four hydroxylated fatty acids produced by Lactobacillus plantaram (3-hydroxydecanoic, 3-hydroxy-5-cis-dodecenoic, 3-(R)-hydroxydodecanoic, and 3-(R)-hydroxytetradecanoic acid) also proved to be relevant in acting against yeasts, with MICs ranging from 10 to 100 μg/mL [133]. However, the mechanisms of how fatty acids inhibit the growth of yeasts are still unclear. It is believed that fatty acids generate the partition of lipid bilayers present on the membranes of fungi, resulting in instability and a loss of permeability control. In these scenarios, fluidity results in the liberation of intracellular proteins, enzymes, and electrolytes, disintegrating the fungal cell [134].

Endophytic bacteria and filamentous fungi are also important sources of bioactive compounds. These microorganisms live within plant tissues, taking their nutrients from this habitat and producing metabolites with some capability of providing protection to the host [135]. In this regard, an endophytic fungus isolated from the leaves of Psidium guajava L., identified as Alternaria tenuissima, demonstrated activity against the human pathogenic yeast C. albicans [136]. The study focused on exploiting the metabolites extracted with ethyl acetate (EA) from the endophytic mold. Different concentrations of the EA extract were analyzed against C. albicans, and a maximum inhibition zone of 14 ± 1.0 mm at a concentration of 10 mg/mL was observed. Moreover, a decrease in the number of colony-forming units (CFUs) of a C. albicans culture treated with this EA extract was observed. The number of CFUs counted suggested that the MIC value of the so-referred extract against this pathogen was 1400 µg/mL. The EA extract from the culture of the endophytic fungus Xylaria sp. PSU-D14, isolated from the leaves of Garcinia dulcis, also exhibited antifungal activity against C. albicans, with a MIC value of 128 µg/mL [137]. From the broth extract of this endophytic fungus, a glucoside derivative, xylarosides A, along with another known compound, sordaricin were isolated. This metabolite demonstrated antifungal activity, against C. albicans, with a MIC value of 32 µg/mL [138].

The alkaloid aspernigrin A, which has been isolated from Cladosporium herbarum (an endophyte of the Cynodondactylon), is also effective in acting against C. albicans. Zhang et al. [139] showed that this compound is able to inhibit this yeast’s growth with a MIC value of 75.0 μg/mL. Another compound, cryptocandin A, isolated from the endophytic fungus Cryptosporiopsis quercina, was characterized as a lipopeptide with antifungal properties. Cryptocandin A demonstrated activity against some important fungal pathogens of humans and showed MIC values of 0.03–0.07 μg/mL against isolates of C. albicans, Trichophyton mentagrophytes, and Trichophyton rubrum. The most significant result was against C. albicans, with a MIC value of 0.03 μg/mL. This value was very similar to the values obtained with amphotericin B, a well-known antifungal agent used in most cases—clinically expressing the potential of this new substance. Cryptocandin A contains some hydroxylated amino acids, for example, the 3-hydroxy-4-hydroxymethyl proline, which can contribute to its activity [140]. The main bioactive compounds produced by bacteria and molds against yeasts are summarized in

Table 2. Relation between microorganisms, biocompounds produced, targets, and MIC.

Producing Microorganism	Biocompound	Target Yeast Tested	MIC	Reference
Alternaria tenuissima	-	Candida albicans	1400 µg/mL	[136]
Cladosporium herbarum	Alkaloid-aspernigrin A	Candida albicans	75.0 μg/mL	[139]
Cryptosporiopsis quercina	Lipopeptide cryptocandin A	Candida albicans	0.03 μg/mL	[140]
Cryptosporiopsis quercina	Lipopeptide cryptocandin A	Cryptococcus neoformans	-	[140]
Pestalotiopsis foedan	Isobenzofuranone, Pestaphtalides A	Candida albicans	-	[141]
Xylaria sp.	-	Candida albicans	128 µg/mL	[137]
Xylaria sp.	Sordaricin	Candida albicans	32 µg/mL	[138]
Nannocystis	-	Candida albicans and Pichia anomala	-	[142]
Streptomyces sp.	Cyclo(l-leucyl–l-prolyl)	Candida albicans; Candida glabrata; Candida tropicalis;	32 mg/mL 16 mg/mL 8 mg/mL	[143]
Streptomyces sp.	Cyclo(l-phenylalanyl–l-prolyl)	Candida albicans; Candida glabrata; Candida tropicalis;	64 mg/mL 256 mg/mL 32 mg/mL	[143]
Lactibacillus plantaram	3-hydroxy tetradecanoic acid	Kluyveromyces marxianus	10 μg/mL	[133]
Lactibacillus plantaram	3-hydroxy dodecanoic acid	Kluyveromyces marxianus	25 μg/mL	[133]
Lactibacillus plantaram	3-hydroxy undecanoic acid	Kluyveromyces marxianus	25 μg/mL	[133]
Mycosphaerella sp.	2-amino-3,4-dihydroxy-2-25 (hydroxymethyl)-14-oxo-6,12-eicosenoic acid	Cryptococcus neoformans	1.95–7.82 μM	[144]
Mycosphaerella sp.	Myriocin	Cryptococcus neoformans	0.48–1.95 μM	[144]

MIC: minimum inhibitory concentration; -: concentration not mentioned.

.

Candida auris and Cryptococcus neoformans: Two Emerging Pathogenic Yeasts to Be Tackled with Bioactive Compounds

Among the yeast pathogens that infect humans and cause them diseases, Candida auris and Cryptococcus neoformans are definitely worth noting. Recently, the World Health Organization (WHO) considered both species critically harmful to human health, showing resistance against antifungals and being responsible for increased death rates [145].

Cryptococcus neoformans infect the lungs, causing pulmonary infections such as pneumonia that can happen before dissemination to the central nervous system [146]. Cryptococcosis and meningitis, caused by C. neoformans, are responsible for a considerable number of deaths in the world [147]. This yeast usually infects immunocompromised people, and it has recently gained attention during the COVID-19 pandemic in patients with SARS-CoV-2 co-infection, especially in those over sixty years old [148].

Two compounds, eicosanoic acid and myriocin, isolated from the endophytic fungus Mycosphaerella sp. (found to be associated with the plant Eugenia bimarginata) were tested against Cr. neoformans, and their MIC values were obtained by microdilution assays. For eicosanoic acid, the MIC values ranged from 1.95 to 7.82 μM, and myriocin’s MIC values ranged from 0.48 to 1.95 μM [144]. Other compounds that demonstrated activity against this yeast were isolated from endophytic fungi associated with Acanthus ilicifolius var. xiamenensis [149] and Tripterigeum wilfordii [140], demonstrating the potential of endophytic molds as a source of novel compounds with bioactivity against yeasts.

Candida auris was described as a pathogenic yeast during the last decade. Over 25% of its strains naturally present multidrug resistance against antifungal compounds, which may be related to their surface stability. Also, the widespread presence of C. auris worldwide may facilitate its contact with different compounds and thus increase its resistance [150]. The first report was made in 2009, receiving attention from medical journals and also mass media [151,152,153].

Despite the search for biocompounds that could inhibit the propagation of C. auris, it remains a challenge to obtain them from sources such as filamentous fungi. However, studies report possible synthetic or semisynthetic options, from natural sources, that can inhibit C. auris growth [154,155].

4. Using Yeasts as “Microbial Cell Factories” of Bioactive Compounds

Yeasts are widely known for producing natural compounds and have been applied in several industrial sectors, such as the production of proteins, pigments, vitamins, fuels, beverages, and foods [156]. Saccharomyces cerevisiae, the main microorganism used in alcoholic fermentation environments, in addition to producing ethanol, has been associated with the release of secondary metabolites responsible for flavors and aromas; for example, in the production of wine, sake, and cachaça, different S. cerevisiae strains have already been correlated with the formation of acetate esters, ethyl esters, acids and higher alcohols [157]. Nevertheless, although S. cerevisiae plays a central role in winemaking, yeasts such as Torulaspora delbrueckii have been gaining prominence due to their greater release of aromatic compounds and the greater intensity of the color of the final product [158,159]. In table olive production, various yeasts, through the generation of metabolites such as glycerol, ethanol, higher alcohols, esters, and other volatile compounds, shape the texture and flavor characteristics of the final product [160]. In addition to food and beverage production, platform molecules have also been explored in yeast-based bioprocesses; for example, isolates of S. cerevisiae and Yarrowia lipolytica have already been tested for the production of succinic acid—an important industrial chemical that is currently produced by petrochemical processing [161,162].

As stated before, bioactive compounds are produced by organisms such as plants, fungi, and bacteria [14,15]. These metabolites can be isolated directly from their natural producers, from fungal and bacterial cultures, or from their accumulation in plant biomass. However, extracting these compounds through direct isolation is difficult, especially in plants with low concentrations, which makes the natural extraction of these products environmentally destructive and economically unsustainable [163]. For example, the natural concentration of taxol (a diterpene alkaloid used against several types of cancer) is, on average, 0.01–0.06% of the dry weight of the plant Taxus spp. [164]; however, the estimated amount of purified taxol needed to treat 500 cancer patients is 1 kg, which is equivalent to about 10 tons of bark or the felling of 700 trees [165]. In addition to this low concentration, seasonal variability and government policies can also influence the productivity of plant crops and, consequently, the extraction of the compounds of interest [166]. Some of these natural products can also be obtained through chemical synthesis, but their structural complexity generally makes this synthetic route difficult, and there are often toxic agents present as byproducts [167,168]—among them, especially, are solvents such as benzene, dichloromethane, and chloroform [169]. In this context, producing bioactive compounds using genetically modified microorganisms represents an advantageous technological alternative in environmental, economic, and health terms.

With the recent advances in genetic engineering (or genome editing, to use today’s more popular term), bioprocesses based on recombinant microorganisms have become increasingly sought after as an alternative to traditional techniques of direct isolation of compounds or their chemical synthesis [170,171]. Fast- and controlled-growth microorganisms, such as the bacterium E. coli and the yeast S. cerevisiae, are already widely used as platform cells to produce high levels of chemicals of human interest, such as biofuels and proteins [172,173]. The yeast S. cerevisiae, in particular, has been shown to be suitable to host several heterologous biosynthetic pathways, given the following traits: (i) being a model organism for molecular biology research, (ii) being the first eukaryotic organism to have its genome completely sequenced, (iii) having the properties of robust growth and high productivity, (iv) being the best-studied organism among lower eukaryotes, and (v) having a wide variety of commercially available and ready-made mutant strains for laboratory use [174,175]. In addition, yeasts have a eukaryotic subcellular organization capable of post-translationally processing many proteins from more complex eukaryotic organisms [176,177]. These factors have led to the use of S. cerevisiae strains as veritable versatile biofactories.

Arguably, S. cerevisiae has a long history of producing compounds of commercial interest. This yeast is widely used to produce fermented foods, beverages, and biofuels, as well as pharmaceutical products such as recombinant human insulin—which since the early 1980s has been massively produced using this microorganism [178]—and the industrial-scale production of the antimalarial precursor artemisinic acid [179]. With the advent of the CRISPR-Cas9 genome editing system [180], rapid and precise genetic manipulation has favored the insertion of a myriad of new biosynthetic pathways in S. cerevisiae, many of which provide products originally found and produced by plants [181]. With CRISPR-Cas9, strains of S. cerevisiae have already been developed to produce the opioids thebaine and hydrocodone from sugars. To achieve this purpose, 21 enzymes were expressed for producing the former and 23 were expressed for producing the latter, with genes from plants, mammals, and bacteria [182]. This yeast has also been modified to produce resveratrol directly from glucose or ethanol [183]. The CRISPR-Cas9 tool was also used to express the last five steps of the carotenoid biosynthetic pathway in S. cerevisiae [184]. More recently, S. cerevisiae has been designed as a platform for the production of daidzein, a key chemical for isoflavonoid biosynthesis [185].

During the last decade, the genomes of several nonconventional yeast species have also been sequenced and made available in databases [186]. In many cases, the interest in new yeasts exists due to the difficulties in using S. cerevisiae as a biofactory, especially when exploring alternative substrates. For example, this yeast mainly uses hexoses as carbon sources, which is an obstacle in the fermentation of sustainable substrates, such as lignocellulosic hydrolysates, which contain, in addition to hexoses, a high concentration of pentoses [187]. Because of this, nonconventional yeasts have been developed as alternative platforms for the production of several recombinant proteins. In fact, the species Y. lipolytica, Kluyveromyces lactis, K. marxianus, H. polymorpha, C. tropicalis and K. phaffii have successfully fulfilled this role (

Table 3. Examples of biocompounds produced by engineered non-Saccharomyces yeasts.

Yeast	Compound	Application	Reference
Yarrowia lipolytica	β-carotene	Antioxidant	[190]
	Linalool	Vitamin precursor, antifungal, antimicrobial and fragrance fixative	[189]
	Lycopene	Antioxidant	[191]
Kluyveromyces lactis	Brazzein	Sweetener compound	[193]
Kluyveromyces marxianus	Astaxanthin	Antioxidant	[194]
Hansenula polymorpha	Gamma-linolenic acid	Anti-inflammatory and imunorregulatory	[195]
Candida tropicalis	α-Humulene	Anti-inflammatory	[192]
Komagataella phaffii (formerly Pichia pastoris)	Lycopene	Antioxidant	[196]
	Lovastatin	Antihypercolesterolemia	[197]
	Δ9-Tetrahydrocannabinol	Compound with analgesic properties	[198]
	Astaxanthin	Antioxidant	[199]
	Citrinin	Antibiotic and antifungal	[200]

). This success is mainly due to their (i) capacity to grow under inhibitory conditions, (ii) tolerance to high temperatures, (iii) ability to use different carbon sources, and (iv) high potential for extracellular protein secretion [188]. Among these yeasts, the oleaginous Y. lipolytica has been genetically modified to produce some bioactive compounds, such as linalool [189], β-carotene [190], and lycopene [191]. Recently, the oleaginous C. tropicalis was modified to produce α-Humulene, a terpenoid with multiple pharmacological activities [192].

Concomitantly with genome editing, evolutionary engineering has also proven to be a resource for directing and selecting the best microbial cell factories. During stages of adaptive evolution, the microorganism of choice is grown for an extended period of time (which can be weeks or even years) under specific conditions that push it towards a desired trait. Afterward, whether or not a more fit and enriched strain with new advantageous mutations has been developed is checked [201,202]. Adaptive laboratory evolution has already been performed, for example, to increase carotenoid production in a recombinant S. cerevisiae strain through its prolonged exposure to hydrogen peroxide stress, since carotenoids have antioxidant properties [203]. In another study, buoyancy was used to select mutants of a recombinant Y. lipolytica strain with a higher ability to accumulate lipids [204]—once lipids reduced cell density.

In short, the ability to heterologously produce bioactive compounds in yeast demonstrates advantages over plant extraction and chemical synthesis. In addition to reducing environmental damage (caused by the need for large areas of cultivation for plant extraction), the biosynthesis of complex molecules can be achieved through new molecular biology techniques and increased knowledge about the genetic characteristics of yeasts. Furthermore, compounds that normally exist at low natural levels can be re-engineered in microbial biofactories for increased productivity [205]. Finally, the vast and unexplored biodiversity of nonconventional yeasts can further optimize the production of bioactive compounds, especially considering the efficient use of renewable and low-cost raw materials as substrates for these biotransformations.

5. Final Considerations

Although higher eukaryotes may produce bioactive compounds, the great demand for these molecules makes it unfeasible and unsustainable to rely on plants and animals for this aim on industrial scale. On the other hand, microbial cell factories perfectly fulfill this role; in this regard, yeasts stand out as the most successful microorganism employed in bioprocesses [11].

Despite Saccharomyces yeasts being seen as the first option for biotechnological purposes until the late 20th century, many studies have recently highlighted the potential applications of unconventional yeasts in the pharmaceutical and food production sectors, as we have discussed above. In this sense, the recent discoveries of new valuable microorganisms should call attention to the social, economic, and environmental importance of biodiversity and bioprospection.

Thus, in the light of circular economy approaches and the one health concept, new public policies and international laws are imperative to induce (i) the development of alternatives to the current environment-impacting production chains, (ii) optimization strategies to increase the production of bioactive compounds from residual feedstocks using yeasts as microbial cell factories, and (iii) the substitution of synthetic pesticides and plant growth hormones in food production systems.