Toward Sustainable Broiler Production: Evaluating Microbial Protein as Supplementation for Conventional Feed Proteins

Abstract

The increasing demand for sustainable poultry production has urged the exploration of alternative feed strategies supporting animal performance and environmental goals. The first section outlines the protein requirements in broiler nutrition (19–25% crude protein) and the physiological importance of balanced amino acid profiles. Vegetal conventional protein sources are discussed in terms of their nutritional value (12.7–20.1 MJ/kg), limitations (antinutritional factors), and availability. Emerging trends in broiler nutrition highlight the integration of supplements and the need for innovative feed solutions as support for the improvement in broiler body weight and feed efficiency increase. Microbial protein sources: yeast biomass (41–60% of 100 g dry weight), microbial mixed cultures (32–76% of 100 g dry weight), and beer by-products, such as brewer’s spent yeast (43–52% of 100 g dry weight), offer promising nutritional profiles, rich in bioactive compounds (vitamin B complex, minerals, enzymes, and antioxidants), and may contribute to improved gut health, immunity, and feed efficiency when used as dietary supplements. The review also addresses the regulatory and safety considerations associated with the use of microbial protein in animal feed, emphasizing EU legislation and standards. Finally, recent findings on the impact of microbial protein supplementation on broiler growth performance, carcass traits, and overall health status are discussed. This review supports the inclusion of microbial protein sources as valuable co-nutrients that complement conventional feed proteins, contributing to more resilient and sustainable broiler production and broiler meat products.

1. Introduction

Feeding a growing population in a world of limited resources has become a major challenge, especially in livestock sectors such as poultry, where the demand for high-quality protein continues to grow [1]. In this context, the search for reliable, efficient and sustainable animal feed sources is more demanding than ever. Conventional feed ingredients such as soybean meal [2] have long been the mainstay of poultry diets, but their dependence on arable land, environmental impacts and volatile market prices raise serious concerns for long-term sustainability and novel development is urgent [3].

One promising avenue is the use of microbially derived feed ingredients, often referred to as microbial bioproducts. Microbial bioproducts include a diverse group of high-protein microbial biomass products derived from microorganisms such as bacteria, yeasts, fungi and microalgae [4]. Various microorganisms can be cultivated using renewable, cost-effective substrates, including agro-industrial by-products, making them not only nutritionally valuable but also environmentally responsible [5].

Among all microbial sources, yeasts and mixed microbial cultures are of particular interest due to their favorable nutritional profiles, potential probiotic effects, and compatibility with circular economy principles [6]. At the same time, incorporating spent brewer’s yeast or other fermentation residues into feed [7] not only improves the nutritional content of the diet, but also contributes to waste minimization and resource recovery in the food industry [8]. Before being widely used in broiler systems, microbial protein sources highlight the current limitations in broiler feeding, such as nucleic acid content (5–15% in yeast), nutrient digestibility [9], different amino acid profiles [10], also might present mycotoxins or toxic metabolites [11,12,13,14].

Due to its low cost and fast production cycle, poultry meat has long been valued for its commercial viability. The growth performance and carcass yield of broiler chickens have been significantly improved and supported by genetic and nutritional advances, which have also led to a notable decrease in carcass fat deposits.

Broiler meat is currently the most widely consumed source of animal protein worldwide [15,16]. Beyond its nutritional and gastronomic advantages, poultry production serves as an economic driver, supporting employment, income generation, and food security in numerous countries [17]. Poultry meat consumption has gained substantial economic relevance in over 50 countries, reflecting both its accessibility and global consumer preference [18]. Globally, poultry meat aligned with beef in 2023 to become the second-most-consumed meat, with pork retaining its position as the leading meat source [19,20]. In terms of market dynamics, poultry production is the fastest-growing segment of the livestock sector [21]. This trend enables producers and distributors to forecast demand with increasing precision. Within the European Union (EU), poultry meat holds strategic importance as the EU functions simultaneously as a major producer, consumer, and exporter [22]. Rising demand for affordable animal protein continues to drive consumption across both domestic and international markets [23]. Romania, in particular, occupies sixth position among EU member states in terms of broiler chicken numbers, slaughter volume, and carcass weights [24,25]. The avian meat sector in Romania has shown consistent growth in production and performance indicators. Recently, approximately 321.79 million birds were slaughtered, contributing to a total national output of 510.689 metric tons (MT) of poultry meat, with chicken accounting for 94.9% of this volume [25]. Over 56% of Romania’s chicken meat is produced in the southern region of the country, which also represents the country’s most significant grain-producing area [26,27,28]. The northeastern region contributed approximately 18% of national production in the same year [25]. In the EU, the increasing cost of food and energy is anticipated to further shift consumer preferences toward poultry meat, as it remains more affordable compared to other animal proteins [29]. In addition, by using high protein microorganisms, also recovering by-products such as brewer’s spent yeast not only supports the circular economy, but also might contribute to environmental safety. This review aims to consolidate current information and identify knowledge gaps in the field of microbial protein sources, supporting their advancement as sustainable and functional alternatives in broiler nutrition.

2. Protein Profile in Poultry Diets

In modern, intensive broiler production systems, feed energy levels exceed 2850–3000 kcal metabolizable energy (ME/kg) [30]. The relationship between energy content and other nutrients is important, given that feed intake is influenced not only by the physiological requirements of broilers, but also by feed energy concentration and ambient temperature [31]. As energy density and ambient temperature increase, voluntary feed intake tends to decrease. Broilers have a high requirement for dietary protein, as their rapid growth is mainly determined by protein intake. Of all nutrients, an optimal intake of essential amino acids is the determining factor for efficient growth and muscle mass accumulation. This has led to extensive research on the specific protein and amino acid requirements of broiler chickens. The concept of “ideal protein” has emerged to describe a protein source with an amino acid profile that precisely meets the needs of the bird, without excess or deficiency [32]. Therefore, pursuing a balanced amino acid profile [33] is more important than focusing on crude protein content [34] in order to tailor the perfect diet for broiler chickens. In practice, when it comes to achieving balanced diets for broiler chickens, one approach is to include high biological value amino acids of synthetic origin to supplement the crude protein component.

The ideal amino acid ratio for broilers, typically expressed relative to lysine (100% lysine, 90% total sulfur amino acids), serves as a reference model in diet formulation [30]. However, despite increasing interest in alternative and sustainable protein sources, such as microbial protein sources, the application of the ideal protein concept in assessing their suitability remains limited. This gap highlights the need for standardized evaluation protocols that incorporate ideal amino acid profiles when determining the nutritional value of non-conventional protein ingredients in poultry diets.

Thus, the metabolic effects of the protein in diets are various. In broilers’ nutrition, it is necessary to consume more protein in order to enhance the protein deposition rather than to maximize the live body weight increase [35]. It is noteworthy that there is an inverse relationship between protein and lipid metabolism; fat deposition tends to decrease as protein gain increases [36]. In general, a 1% increase in the dietary protein content (equal to 10 g/kg) [37] generates the carcass lipid content to mitigate by 7–13 g/kg, and the feed consumption to decrease by roughly less than 3% [38]. Moreover, essential amino acids are required for proper nutrition, the majority of those are first limiting, such as methionine, cysteine, lysine, and threonine [39]. On the other hand, the growth performance, muscle and skeletal development, and the manifestation of genetic potential can all be severely hampered by deficiencies in these amino acids. Furthermore, these deficiencies and abnormalities may impair anabolic processes, hence compromising the feed efficiency and overall productivity [40]. Additionally, protein-rich feeding ingredients, such as soybean meal, might result in an excess of amino acids, especially the ones that are higher than that required by the chick’s metabolism. Usually, when the chick receives amino acids that exceed its requirements, it goes though catabolic processes that might cause the level of nitrogen excretion to increase [41]. Uncontrolled excess nitrogen is considered a nitrogenous waste product, mainly as nitrates, and is closely linked to environmental pollution [42].

When evaluating novel protein sources, such as yeasts, they are scrutinized for their valuable advantages for industrial applications, including rapid growth rates, the ability to utilize a wide range of carbon-rich organic waste substrates, and cost-effective production and harvesting processes that require minimal human or technological intervention. These attributes make yeasts particularly well-suited for large-scale protein production and position them as competitive candidates within the commercial protein market. The plant-based sector currently serves as the primary source for natural proteins for human and livestock nutrition. However, its productivity is increasingly constrained by factors such as declining soil fertility and the adverse effects of climate change, resulting in supply chain disruptions and significant economic repercussions across protein-dependent sectors, including food, feed, pharmaceuticals, and cosmetics. In this context, microbial-derived proteins are gaining attention as a promising alternative. Although still under development, microbial protein production offers notable advantages, including higher crude protein values, and reduced production time, making it a viable and sustainable substitute for conventional plant-based sources in the near future.

It is mandatory to tailor the supplemented microbial-based biopreparation diets with a balanced nutraceutical profile in order to support the chick’s optimal growth and development as well as to maintain general health status. Concerning the unique nutritional requirements [41,43] of each livestock species, it is important to evaluate if the dietary formulation meets the suitable amino acid composition [44]. Aligning the dietary amino acid profile and composition with the optimal needs for maintenance, growth and productivity is becoming more and more important in precision feeding strategies [45].

Lysine is generally the first limiting amino acid in the conventional feeding formula for monogastric species [46,47]. It has an important role in the synthesis of body proteins, peptides, and various non-peptide compounds. Any surplus of lysine is metabolized for energy [48]. For poultry species, lysine also holds a high nutritional significance, but sulfur-containing amino acids, specifically methionine and cysteine, are considered the primary limiting nutritional factors [49]. These amino acids are essential for feather development immediately post-hatch and play a significant role in supporting the endocrine system. According to recent studies [50,51], feed formulas for monogastric species are commonly balanced using synthetic amino acids such as L-lysine and DL-methionine, which are obtained through microbial biosynthesis. Digestibility is another important factor influencing protein ingredient quality. Nutritional outcomes can be greatly impacted by the degree to which amino acids are broken down and absorbed in the chick gut. It has been demonstrated that brewer’s spent yeasts (BSY) and inactive or hydrolyzed yeast biomass (HYB) contain significant amounts of nucleic acids (ranging from 5 to 15%) [52], which can limit their use in monogastric diets. Moreover, the consumption of high levels of nucleic acid in diets, in both human and animal nutrition, are often related to potential health issues, such as gout and kidney failure [52].

High levels of spent yeast in poultry diets have been associated with adverse health effects [53,54]. Moreover, microbial protein biomass often presents limitations in quality due to the low concentration of one or more essential amino acids, rendering them limiting and thus impacting the protein’s nutritional adequacy.

3. Conventional Protein Sources in Broiler Diets: Nutritional Profiles and Utilization

Soybean meal, lentils, chickpeas, canola meal and sunflower meal are examples of plant-based protein resources. Sources such as soybean meal, sunflower meal, and canola meal are represented in Figure 1. Generally, conventional plant-based sources offer a variety of vital minerals, vitamins and other nutrients in addition to proteins. However, the protein constituents in broiler diets need to be balanced in precise amounts and supplemented with other protein sources in order to attain the correct protein concentration and amino acid composition. This method lessens the possibility of production and health problems brought on by consuming insufficient or excessive protein.

3.1. Soybean Meal

Soybean meal is recognized for its high biological value, offering a balanced amino acid profile comparable to that of animal-derived proteins. The limiting amino acid present in soybean meal is methionine, which is typically supplemented to balance the lysine–methionine ratio, thus optimizing protein utilization [55]. Soybean meal provides approximately 80% of the essential amino acid requirements for broilers [56], making it an indispensable ingredient in poultry feed. The digestibility of amino acids from soybean meal is notably high, around 91%, which enhances its nutritional efficiency. However, despite its advantages, soybean meal requires supplementation with methionine due to its deficiency in this amino acid. It also contains a variety of other essential nutrients, including sodium, choline, riboflavin, folic acid, niacin, pantothenic acid, and thiamine [57]. On a global scale, soybean meal accounts for two-thirds of the total plant-based protein feed production, with major exporters such as Argentina (33.5 million metric tons) and Brazil (30 million metric tons) leading the market [58]. Within the European Union, soybean meal represented over 60% of the total protein used in animal feed in 2023, amounting to a total of 24 million metric tons [59]. Soybean meal is categorized into conventional soybean meal (43–44% protein content) and high-protein soybean meal (47–49% protein content), both offering high biological value [60]. Nevertheless, soybean meal has certain limitations. It contains oligosaccharides such as raffinose and stachyose, which are indigestible by poultry, and these compounds have been shown to reduce metabolizable energy, fiber digestion, and transit time [61]. Additionally, the high cost and global availability of soybean meal present economic challenges, especially when local sources are unavailable. Despite its inclusion typically not exceeding 30% in broiler diets, it constitutes approximately 40–50% of the total cost of compounded feed production [62]. Moreover, the environmental impact of soybean cultivation, particularly deforestation in the Amazon region of Brazil, raises socio-economic concerns [63].

3.2. Sunflower Meal

Sunflower, a widely cultivated crop, is adaptable to a variety of climatic and soil conditions [64,65]. The residual meal, when appropriately processed, offers a protein profile comparable to soybean meal, though it is generally lower in essential amino acids. The nutrient composition of sunflower meal varies based on seed quality and the extraction method used. Efficient decortication before oil extraction significantly impacts the quality of the meal, with protein content typically around 40% and fiber content under 13%. Sunflower meal contains fewer antinutritional factors compared to soybean meal and is more resistant to contamination. However, the presence of over 20% crude fiber poses a limitation in broiler feeding, particularly for young birds, due to their small digestive tracts and high protein requirements [66]. Moreover, sunflower meal contains high levels of chlorogenic acid, a phenolic compound known to inhibit digestive enzymes such as trypsin, chymotrypsin, amylase, and lipase [67]. The typical composition of sunflower meal is 60–65% seed kernels and 35–40% hulls, with protein content ranging from 30% to 34%, fiber at 20–25%, and lignin at 8–10% [68]. Compared to soybean meal, sunflower meal has certain nutritional drawbacks, including a high hull content, a relatively high iron content, and a lower metabolizable energy value and lysine content.

3.3. Canola Meal

Canola meal, a byproduct of rapeseed oil extraction, ranks second globally in terms of production after soybean meal [2]. The nutritional quality of canola meal is influenced by both the type of cultivar and the oil extraction method used. The optimal conditioning temperature for canola seeds during oil extraction ranges from 100 °C to 105 °C, for 15–20 min [69]. Excessive temperatures during processing can reduce the digestibility of essential amino acids in the meal when used in monogastric animal feeds. Canola meal is notable for its high calcium and phosphorus content compared to soybean meal. However, phosphorus in canola meal is primarily present in the phytate form (65%), rendering it less bioavailable [70]. Additionally, canola meal contains significant levels of sulfur (approximately 1.1%), which can lead to leg abnormalities in broilers [71].

3.4. Lentils

Lentils (Lens culinaris) occasionally become available to the broiler-feed industry, at times when these grains are downgraded qualitatively for human consumption [72] (frost injury, discoloration or deterioration of the grain). These grains present no problem when used as feed for chickens of all physiological stages. The interest in the use of lentil grains in broiler feed is justified primarily by their relatively high protein content [73] (25–29.1% per kg dry matter), their essential amino acid profile (especially lysine, up 6.5–7 g/100 g protein), but also by their low antinutritional factor level content, when compared to soybean, also by their high energy value (up to 12.7 MJ metabolizable energy/kg) [74]. Lentils are also a fairly rich source of minerals (calcium, phosphorus, assimilable iron, zinc, and selenium) and vitamins (thiamine, riboflavin and niacin). There are recent studies regarding lentil dietary inclusion in the levels of 1:4, lentils with soybean meal on broiler starter phase, and over 400 g/kg feed for the broiler-finisher phase [75].

3.5. Chickpea

Chickpea (Cicer arietinum L.) is an annual leguminous plant, characterized by high drought resistance, as the crop is well adapted to semi-arid soil and climatic conditions, with chickpea-generated soil aeration, structured and weed-free [76]. The plant’s external appearance differs from that of peas in the shape of its leaves and pod (which usually encloses two or three pods, rarely a single pod). The use of chickpeas in poultry feed is justified primarily by their relatively high protein content (26.1% per kg dry matter), their essential amino acid profile (especially the limitative ones) [77], but most of interest is due to their low antinutritional factor content compared to soybean (Glycine max L.), and their energy value (12.5 MJ metabolizable energy/kg) [78]. Chickpeas are also a fairly rich source of mineral salts (phosphorus, potassium, magnesium, calcium, iron) and also the B vitamin complex. Recent findings support inclusion of chickpea levels up to 200 g/kg feed, in the broiler-finisher phase [79] (

Table 1. Comparative protein quality of high protein vegetal resources.

Item	SBM	SFM	CM	Lentils	Chickpea
CP%	44–49	34–44	36.5	26.7	26.1
ME [MJ/kg]	20.1	12.7	18.6	12.7	12.5
Amino acid content %
Lysine	2.74	1.18	2.13	1.81	1.84
Methionine	0.6	0.72	0.7	0.79	1.14
Cysteine	0.63	0.55	0.82		0.15
Threonine	1.72	1.21	1.54	1.01	1.22
Tryptophan	0.59	0.45	0.48	-	-
Arginine	3.28	2.68	2.38	2.01	1.78
Glycine	1.86	1.92	1.77	0.92	0.73
Serine	2.25	1.40	1.44	1.25	0.97
Histidine	1.17	0.82	1.22	-	2
Isoleucine	2.13	1.47	1.25	0.91	1.2
Leucine	3.4	2.12	2.22	1.92	1.83
Phenylalanine	2.22	1.50	1.46	1.24	1.39
Tyrosine	1.62	0.81	0.9	0.63	0.71
Valine	2.19	1.78	1.78	1.05	1.15
References	[80]	[80]	[81]	[75]	[79]

SBM—soybean meal; SFM—sunflower meal; CM—canola meal, CP—crude protein; ME—metabolizable energy.

).

4. Trends in Poultry Nutrition: Challenges and Innovations

Recent European Union regulations regarding poultry farming systems, feed quality, and the use of nutritional additives have highlighted the need for the identification and incorporation of alternative feed ingredients. The regulatory changes aim to improve animal welfare and ensure the production of “safe” food products, including meat and eggs, for human consumption. Previous research has demonstrated that yeast biomass and soybean meal supplemented with synthetic methionine and organic selenium can effectively replace animal-derived ingredients in poultry diets [82].

4.1. Protein Bioproducts: Role and Application in Poultry Nutrition

Amino acids hold physiological roles in the avian organism. Following the absorption pathway, amino acids are metabolized to synthesize proteins, which are subsequently utilized in the formation of various body tissues. The recommended levels of essential amino acids in poultry nutrition can vary based on factors such as the specific hybrid of poultry [41,42,43], the growth technologies, or maintenance systems in place. Extensive research has been conducted on the incorporation of synthetic amino acids into poultry diets. Proper supplementation with synthetic amino acids has the potential to enhance the overall amino acid balance, while simultaneously reducing the crude protein content in poultry feed [79]. This approach not only improves dietary efficiency but also contributes to a more sustainable and cost-effective feeding strategy [83]. The use of synthetic amino acids in the formulation of compound feed reduces nitrogen losses in protein metabolism, thereby decreasing nitrogen excretion and improving growth performance in poultry [43]. Additionally, in poultry diets, it is necessary to balance amino acids to minimize energy loss, which could otherwise be redirected toward fat synthesis [84]. There is evidence that digestibility and protein metabolism are affected by the health status of the birds (

Table 2. Broiler protein metabolism affected by health status.

Birds’ Health Status	Potential Effects on Digestibility	Protein Metabolism	References
Boilers with homeostatic status	Efficient digestion and nutrient absorption with optimal enzymatic activity and balanced gut flora.	High protein synthesis rates, efficient amino acid utilization, normal nitrogen retention.	[85]
Presence of coccidiosis infection	Could present damaged intestinal lining, reduced nutrient absorption and impaired enzyme secretion.	Increased protein catabolism, reduced growth performance, poor feed conversion.	[86]
Broilers with necrotic enteritis	Might present inflammation of gut mucosa, with decreased villus height and impaired digestion.	Increased muscle breakdown lowered protein deposition, elevated plasma uric acid levels.	[87]
Broilers infected with Clostridium perfringens	Disruption of gut microbiota with increased digesta viscosity and reduced digestibility.	Altered nitrogen metabolism impaired amino acid absorption.	[88]
Forage mycotoxin exposure	It might inhibit the digestive enzymes’ activities and damage the intestinal epithelium surface.	Inhibition of protein synthesis, increased liver stress decreased nitrogen retention.	[89]
Broilers suffering heat stress	Could alter the gut permeability, with direct effects of reducing feed intake; also enzyme denaturation.	Increased protein breakdown, reduced protein accretion, impaired muscle development.	[90]
Subclinical inflammation or stress on broilers	Present as mild reduction in nutrient digestibility with subtle shifts in gut microbiota.	Redirection of amino acids toward immune response, reduced growth rate.	[91]
Use of antibiotics or probiotics in broilers’ nutrition	Might improve nutrient digestibility through the stabilization of gut flora.	Enhanced protein metabolism, improved nitrogen utilization and feed efficiency	[92]

).

4.2. Microbial Protein Bioproducts

In response to global population growth, economic development, and urbanization, meat consumption has risen exponentially over the past 50 years, reaching over 328 million metric tons in 2021 [93]. The UN predicts that the demand for protein will increase by more than 50% by 2050 compared to 2020 levels [86]. However, inflation caused by global conflicts is driving up the price of meat and of grains used for animal feed. Due to rising import prices of these essential products, the EU faces the risk of food insecurity [94]. Furthermore, precision livestock farming ranks among the top two or three contributors to pressing global environmental issues, such as water usage, air pollution, deforestation, and biodiversity loss [95]. Over the years, significant research efforts have been dedicated to improving the microbial biopreparation production process, leading to the adoption of methanol and ethanol derived from petroleum as suitable substrates for microbial protein production [96,97,98]. Renewable sources, particularly food and agricultural by-products such as molasses and whey, as well as industrial waste rich in starch, cellulose, and hemicellulose, have regained importance as preferred substrates for microbial protein production. These agro-renewable substrates offer economic advantages, ecological sustainability, and compatibility with circular economy principles [99,100,101,102]. As a result, major microbial protein projects based on petroleum derivatives as substrates were gradually abandoned in the 1980s in favor of renewable and sustainable alternatives.

However, concerns over the safety of fermentative substrates and rising oil prices led the industry to refocus on renewable sources. This shift reflects a broader trend towards prioritizing environmentally friendly and economically viable approaches in microbial protein production, aligning with the increasing emphasis on sustainability and responsible resource use. Up to 75% of agricultural land is used for animal husbandry and feeding [103], securing only one-third of the global protein supply [104].

Given the limited natural resources of the planet, this is a cause for concern, prompting stakeholders in these sectors to rethink the efficiency of animal protein production. Soybeans, the most widespread protein source, despite being costly, undoubtedly have an unappealing taste and a strong environmental impact, contributing to nearly 20% of tropical deforestation [63].

Microbial protein consumption is familiar, as humans have been consuming microbially derived products (such as beer, bread, yogurt, and cheese) long before fully understanding their contributions to human health and well-being [105,106,107]. The aim of fermentative biotechnologies is to produce valuable organic metabolites. Compared to conventional proteins, these ingredients are used in much smaller quantities with a high degree of purity.

Despite the complex nature of microbial protein production, which is well-defined and innovative, it is still considered underdeveloped, mainly due to microbial diversity. Selecting the appropriate production microorganism is fundamental in any biotechnological process [108], which is why microorganisms used for protein production are chosen based on fermentation requirements. Recent studies highlight multiple examples of microbial protein production (

Table 3. Microbial protein production.

Microorganism	Microorganism Nutrient Substrate	References
Fungi
Aspergillus oryzae	organic	[111]
Aspergillus ochraceus	organic	[112]
Cladosporium cladosporioides	organic	[93]
Monascus ruber	organic	[113]
Penicillium citrinum	organic	[114]
Yeast
Saccharomyces cerevisiae	organic	[115]
Candida utilis	organic	[116]
Kefir sp.	organic	[117]
Kluyveromyces marxianus	organic	[118,119]
Yarrowia lipolytica	organic
Hanseniaspora uvarum	organic	[120]
Bacteria
Rhodopseudomonas blastica	mix	[121]
Escherichia coli	organic	[122]
Bacillus subtilis sp.	organic	[123]
Bacillus cereus	organic	[122]
Corynebacterium ammoniagenes	organic	[124]
Algae
Ulva fasciata	mix	[125]
Laurencia intricate	mix	[126]
Chlorella sorokiniana	mix	[127]
Arthrospira platensis	mix	[128]
Chlorella sp.	mix	[129]

) by heterotrophic bacteria, yet most industrial-scale microbial protein sources have been synthesized by yeasts or fungi [109]. Among the microorganisms used for microbial protein production, microalgae have the highest protein content (61–72%), followed by bacteria (31–81%), yeasts (29–53%), and protists (11–22%) [110].

4.2.1. Yeasts

The biotechnological applications of yeast, a heterogeneous group of eukaryotic fungi, are currently limited to a small number of species, including Candida utilis [130], Kluyveromyces marxianus [131], Yarrowia lipolytica [132], and Pichia pastoris, with Saccharomyces cerevisiae occupying a predominant position [133]. Yeasts have the ability to grow on various substrates, contain a high protein content (45–55% of dry weight), and are rich in B vitamins, making them one of the most commonly used microorganisms [134]. Besides their ability to grow at acidic pH and be easily harvested (due to cell size), yeasts have the essential advantage of being well-known and accepted by consumers, having long been used in traditional fermentation (baking, beer, and wine) [135]. Along with having a lower methionine concentration, when compared with bacteria, yeasts have a higher lysine content [93]. Traditionally, yeast extracts have been made from Saccharomyces cerevisiae, also referred as baker’s or brewer’s yeast. It is also utilized in the production of savory spreadable goods [133].

Recent research suggests that a variety of agro-industrial waste and residues, including wine yeast [136], effluents from oil extraction [137], orange peel residues, and hemi cellulosic residues from sugarcane processing [138], can be used to produce microbial protein using yeasts from Candida genus. However, using Candida can be challenging due to the fact that most of them are still opportunistic in terms of infections; in humans, species such as Candida tropicalis, Candida albicans, Candida galbrata, Candida prapsilosism and Candida krusei [139]. Due to its potential pathogenicity, there is limited literature on the industrial biotechnology of C. krusei. However, C. krusei has a wide range of biotechnological applications, and its presence in many traditional foods, such as dairy products, suggests that it does not secrete mycotoxins in the final fermented product.

Yarrowia lipolytica is a yeast that is phylogenetically distinct from other studied yeast species. The FDA has recognized its metabolites as generally regarded as safe (GRAS), and the European Food Safety Authority (EFSA) approved Yarrowia lipolytica biomass as an innovative novel food in 2019 [140]. Kluyveromyces marxianus is a yeast that utilizes lactose from whey and whey milk [141], making it an excellent candidate for microbial protein production and widely used as a feed microorganism [142].

4.2.2. Mixed Microbial Cultures

The application of mixed microbial cultures has been proposed to enhance biomass yield and improve protein quality [116,143]. However, the interactions between strains during mixed fermentation still require further clarification. The microorganisms Candida tropicalis, Trichoderma koningii, and Aspergillus oryzae have been evaluated for microbial protein production using orange waste. A recent study [111] reported both synergistic and antagonistic effects during mixed fermentation between T. koningii and A. oryzae. The growth of C. tropicalis was inhibited by T. koningii and A. oryzae due to the accumulation of significant amounts of polygalacturonase and carboxymethylcellulase [111].

A yeast mixture consisting of Kluyveromyces lactis and Rhodotorula graminis was shown to be effective for microbial protein production. Mixed yeast cultures exhibit higher productivity in protein yields while simultaneously reducing total organic carbon (TOC) [144]. Various microbial combinations have been suggested for microbial protein production using whey as a substrate, such as Kluyveromyces marxianus and Candida krusei [145], Torulopsis cremoris and Candida utilis [144], and Kluyveromyces marxianus and Saccharomyces cerevisiae [146].

4.2.3. Brewer’s Spent Yeasts

Throughout the entire agri-food supply chain, enormous amounts of waste and valuable by-products are generated [97,99,101]. Inefficient and unsustainable management of these by-products and waste reflects the socio-economic situation of a region. Additionally, environmental issues and policies adopted at the regional level can also have negative impacts. Brewing produces [147] a vast amount of various secondary products. Most of these by-products are commonly used as fertilizers on fields [148], incinerated, or discharged into wastewater as waste. This type of by-product management presents a significant environmental challenge [104] that needs to be addressed through the development of new technologies and the redirection of these biotechnologically important by-products. Minimizing the generation of by-products or reintegrating them into the production process as raw materials can contribute to achieving ecological sustainability. Given that some by-products from the malting and brewing industries are not only nutritious and valuable but also inexpensive and readily available [149], they can be incorporated into various industries focused on the production of food, pharmaceutical, or biotechnological products. The beer production process involves a series of unit operations. Considering that the ratio of beer produced to wastewater is 1:10 [150], it is not surprising that water is the most abundant by-product in the brewing industry. In addition to water, other by-products from brewing include spent grains [151], spent yeast [134], and spent hops/hot wort [152]. Malting production also uses a significant amount of water and generates germinated grains/roots. Regardless of the by-product, there is a need to find new methods to utilize them, ensuring safe disposal into nature or minimizing their occurrence.

After fermentation and maturation, yeast precipitates (Figure 2) in the tank designated for bottom fermentation or rises to the surface during warm fermentation [105]. While most of the yeast can be easily separated after decantation or flotation in the tank, some yeast remains, and this fraction must be removed through centrifugation or filtration [153]. Commercial brewing yeast is inactive and rich in proteins, minerals, vitamins, nitrogen, and enzymes [154]. To prevent enzyme inactivation, the fermentation process must be halted under controlled conditions (T 29–61 °C; t = 12.4–18.1 h) [155]. Spent yeast is bitter and should not be confused with pure brewing yeast found on the market, which is produced under specific controlled conditions [156].

Although brewing yeast can be reused several times in the production process (four to six times), it still generates a significant number of by-products due to the rapid growth and reproduction of its biomass during fermentation [154]. However, waste such as spent brewing yeast and plant residues from the fermentation process are often disposed of by being dumped in fields or incinerated [157]. Recently, several studies were conducted to find biotechnological solutions for this by-product. Spent brewing yeast can replace 50% of the protein in fish feed or be added as a supplement (up to 30%) without negatively affecting growth, development, or meat quality [158]. Furthermore, spent brewing yeast has shown great potential in human nutrition due to its high protein, mineral, and vitamin content. Spent yeast can be processed into concentrates and isolates available commercially in the form of powders, tablets, flakes, or liquids. Liquid commercial spent yeast contains enzymatically degraded yeast for easier digestion, absorption, and utilization. The main challenges arise from protein isolation, as they are rich in nucleic acids, particularly RNA [115]. Therefore, further research focuses on isolating proteins with lower RNA content [159]. Fermentation at low temperatures produces large quantities of crude protein (40–60%) [160]. Spent brewing yeast is a good source of nicotinic acid, cysteine, glycine, and glutamic acid [161]. In addition to the B vitamin complex, one of the most important metals found in spent brewing yeast that affects human health is trivalent chromium [162]. Due to its nutritional properties, spent brewing yeast is often used as a suitable medium for the growth and development of microorganisms. Its applications in microorganism production have significant economic importance due to the development of a branch of the food industry that generates functional foods [115].

5. Regulations and Legal Standards on the Use of Microbial Protein Sources in Animal Nutrition

Despite the growing interest in microbial protein sources as sustainable and efficient alternatives to conventional protein sources, their widespread adoption in animal nutrition faces several challenges. One of the major concerns lies in the high nucleic acid content (5–15%) [115] found in many microbial protein sources [9], which may lead to uric acid accumulation and related health issues, particularly in monogastric animals [115]. Furthermore, some microbial protein sources exhibit lower digestibility due to rigid cell walls or the presence of antinutritional factors, limiting amino acid bioavailability. Additional safety concerns are raised by the potential presence of toxins, endotoxins, or undesirable metabolites produced during microbial fermentation [9,12]. From an economic perspective, the production of microbial protein sources remains relatively expensive, as it requires sterile fermentation environments, specific substrates, controlled aeration, and extensive downstream processing [29]. These technological demands can significantly elevate production costs compared to traditional protein sources such as soybean meal or fishmeal. Furthermore, palatability issues, potential nutritional imbalances, and limited consumer familiarity may hinder broader acceptance in the feed industry. Another important aspect is the regulatory complexity associated with introducing new microbial protein sources into the feed chain. Obtaining authorization involves rigorous safety assessments and compliance with both EU and national regulations, creating potential bottlenecks in innovation and market access. Nonetheless, when produced from renewable substrates or waste streams, microbial protein sources hold considerable promise in contributing to circular economy strategies and reducing the environmental footprint of livestock production [29,136].

The incorporation of microbial protein sources in animal feed must comply with relevant food and agricultural regulations and legal standards, including Regulation (EC) No. 183/2005 [163], Regulation (EC) No. 178/2002 [164], Regulation (EC) No. 767/2009 [165], and Directive 2002/32/EC [166]. These regulations cover food safety standards, labeling requirements, and approvals for food additives Regulation (EU) 2023/915. The use of microbial-derived proteins is supported by EU legislation, specifically Regulation (EU) No. 2017/1017 [167].

The European Food Safety Authority (EFSA) maintains a Qualified Presumption of Safety (QPS) list [168], which identifies microorganisms deemed safe for use in food and animal feed. However, some conflict points are highlighted as follows:

• The EFSA QPS list is based at the species or group level, not the strain level.
• Some species present on the QPS list have beneficial effects, and some potential harmful effects, and this could mask pathogenic potential, especially for genera like Bacillus, Enterococcus, or even certain Lactobacillus species.

Additionally, the EU has a strategic framework for addressing food security, health, and sustainability by 2030, under the “Food 2030” policy initiative [169]. Microbial protein sources are part of the alternative protein resources framework, which aims to contribute to changing diets without negatively impacting the climate by 2030. The EU also funds research into alternative protein sources. Within the Horizon 2020 program, the European Commission invested EUR 70 million in 15 different projects investigating the potential of plant-based proteins, insect proteins, and microbial protein sources. At the national level, the use of protein hydrolysates is authorized according to CE No. 741/2003 [170], which amends the Veterinary Sanitary Norms for preventing, controlling, and eradicating certain transmissible spongiform encephalopathies, as approved by the Ministry of Agriculture, Food, and Forestry Order No. 144/2002. Furthermore, microbial protein sources are included in the catalog of approved feed materials, as outlined by Regulation No. 68/16.01.2013 [171], adopted by the European Commission.

6. Assessment of the Impact of Microbial Protein Alternatives on Poultry Growth Performance and Health

The use of microbial protein sources derived from brewery industry waste offers notable sustainability benefits and economic efficiency (

Table 4. Effects of yeasts * supplemented in broiler nutrition on performance and health status.

References	Level (% in Diet)	Body Weight Gain (g/bird)	Feed Conversion Ratio	Immune Markers/Blood Markers	Other Effects/Remarks
[172]	SC: 0, 25, and 50%, to replace corn gluten meal (CGM)	25% had comparable body weight (BWG), feed intake (FI), and feed conversion ratio (FCR) to the birds fed only CGM.		-	Improved gut morphology
[173]	SC: 20 g/100 kg feed	Significant improvement in body weight, body weight gain, feed efficiency, and carcass yield compared to that of the control.		Antibody titer against Newcastle disease (ND) and infectious bronchitis disease (IBD) was significantly increased in PB and AGP groups.	Optimal performance, improved blood profile
[53]	SC: 0, 0.6, 1 and 1.3 g/kg feed	Having growth-promoting and product-quality-enhancing benefits, when fed up till 0.6 g/kg feed.		-	Yeast supplementation could synergically enhance meat quality attributes and might positively modulate the consumer’s preference, increasing meat moisture, lightness, redness, and decreasing the browning index.
[174]	SC: 2.5, 5.0, 7.5, 10.0 and 12.5 g/kg feed	-		When fed with SC: 7.5 or 10.0 g/kg feed, had improved (p < 0.05) packed cell volume (PCV), hemoglobin (Hb), mean cell hemoglobin (MCH) and mean cell volume (MCV) compared to birds.	Balanced and improved blood profile.
[175]	SC: 500 mg/kg in starter and grower phase, and 250 mg/kg in finisher phase	SC promoted growth during d 15 to 28 (p < 0.05)		Significantly increased the jejunum villus height (VH) and the ratio of villus height to crypt depth (VCR) of jejunum, and decreased the crypt depth (CD) of ileum (p < 0.05).	Stronger performance and overall heath
[175,176]	SC: at a 1 g/kg diet and 2 g/Kg feed	SC group resulted in a better (p < 0.05) feed conversion rate (FCR) than the control group		Meat and liver cholesterol, as well as the cholesterol-to-lipid ratio of meat and liver, were significantly decreased (p < 0.05) in SC.	Boost in growth and health performance
[176]	SC and Lactobacillus acidophillus	Mixed supplementation group resulted in a better (p < 0.05) feed conversion rate (FCR) than the control group		↑ Lactobacillus spp., ↓ E. coli GUT populations.	Improved the gut flora, enhanced gut health
[177]	Baker’s yeast: 0, 0.20%, 0.40% dietary feed	No or lowest supplementation affected negatively (p < 0.05) the average daily feed intake (ADFI), final live weight (FLW), average daily weight gain (ADWG), and FCR compared to those in the group supplemented with 0.40% feed.		↑ PCV, ↑ serum albumin.	There were no significant (p > 0.05) differences in white blood cell, red blood cell, hemoglobin, lymphocyte, heterophils, H/L ratio, platelets, mean corpuscular volume, mean corpuscular hemoglobin concentration, and mean corpuscular hemoglobin values across the dietary groups.
[178]	0, 1, or 2 g/kg feed	-		Increased (p  <  0.05) the ileal villus height-to-crypt depth ratio, and ileal goblet cell density in broiler chickens.	SC enhanced nutrient utilization and augmented intestinal development in broiler chickens.

* Different yeast supplementations: Brewer’s spent yeasts/Saccharomyces cerevisiae (SC)/baker’s yeasts/hydrolysate yeasts, ↑—higher, ↓—lower.

).

Converting beverage production residues into valuable protein sources not only reduces the environmental footprint of the food industry by minimizing waste but also enhances energy efficiency and helps lower greenhouse gas emissions. Moreover, microbial protein sources represent a promising, nutrient-rich alternative to conventional protein sources. Their inclusion in animal diets can lead to improved productivity and overall animal health, ultimately influencing the quality and nutritional value of the final products. Recent studies [172,179] indicate that supplementation with BSY, especially in the range of <2–4% of the diet, consistently improved growth performance in poultry. The body weight gain and feed conversion rates reported in the studies suggest high nutritional availability and digestibility of nutrients derived from BSY [53,172]. These results can be attributed to the high-quality protein content, bioavailable amino acids, B-complex vitamins and functional polysaccharides present in Saccharomyces cerevisiae [172,175,176], which together contribute to better growth and feed efficiency. Such consistency across different inclusion levels reinforces the value of BSY [53] as a sustainable and cost-effective supplement protein source in poultry production systems.

In addition to growth performance effects, BSY exhibits pronounced immunomodulatory and antioxidant properties. Increased levels of immunoglobulins (IgG), serum protein fractions, and antioxidant enzyme activities (SOD) reported in multiple studies reflect enhanced immune competence and attenuation of oxidative stress in broilers [179]. The reduction in malondialdehyde (MDA), a marker of lipid peroxidation, further supports the role of BSY in maintaining cellular integrity and systemic health [172]. These findings highlight the potential of BSY as a functional feed additive that not only supports growth but also strengthens the bird’s defense mechanisms, especially under suboptimal growing conditions.

Several studies in the dataset highlight the beneficial effects of BSY on intestinal morphology and microbial ecology [175,176]. Increased villus height [173] and reduced pathogenic E. coli populations [176], accompanied by increases in beneficial microbes such as Lactobacillus spp. and Firmicutes [175,176], reflect a more stable and efficient intestinal environment. These effects are likely mediated by the prebiotic components of BSY, including mannan-oligosaccharides (MOS) and β-glucans, which are known to modulate the intestinal microbiota and enhance mucosal immunity [174]. The improved intestinal architecture and microbial balance observed in response to BSY inclusion may translate into better nutrient absorption, lower incidence of disease, and improved overall performance.

7. Conclusions

This review underlines the multifaceted potential of microbial protein sources, particularly those derived from yeasts and fungi, as viable alternatives to traditional feed components. Production via fermentation processes and subsequent biomass recovery exemplifies a resource-efficient and environmentally conscious approach aligned with circular economy principles. Yeasts and mixed microbial cultures emerge as particularly promising bioresources due to their rich protein content, functional bioactive compounds, and potential probiotic properties. Moreover, the valorization of agro-industrial residues, especially brewery by-products and spent yeast, not only enhances sustainability but also contributes to waste reduction and feed cost optimization. Important parameters such as amino acid composition, protein digestibility, and bioavailability continue to be decisive in determining the efficacy of microbial protein sources in broiler diets. Microbial protein offers a concrete solution to sustainability challenges in animal nutrition by reducing reliance on conventional protein sources such as soybean meal and fishmeal, which are associated with deforestation, overfishing, and high water and land use. Microbial protein sources can be produced in controlled environments using significantly less land and water, while also utilizing renewable carbon sources or waste substrates from agro-industrial processes. This not only lowers the environmental footprint of feed production but also provides a consistent and scalable protein source that is less affected by seasonal variability or climate change. Furthermore, the closed-loop nature of microbial protein production aligns with global climate and sustainability goals, enabling the livestock industry to reduce greenhouse gas emissions and move towards net-zero targets.

By consolidating current knowledge and identifying critical research gaps, this review advocates for the integration of microbial protein sources as sustainable, functional feed components capable of meeting the evolving demands of modern poultry products. Furthermore, experimental research regarding the testing of novel microbial protein biopreparation formulations and the microbial protein biopreparation effects on broiler health and growth performance is needed. In addition, specific directions for research emerge due to the need for knowledge (precision fermentation and microbiome interactions) in order to achieve a better pathway to obtaining microbial protein sources.