Probiotics for Animals: Direct-Fed Microbials, Strains, Spores and Evidence

Probiotics for animals, called direct-fed microbials in the US, are live bacteria, spores or yeasts added to feed or water, and the EU authorises them strain by strain as zootechnical additives, mostly in functional group 4b gut flora stabilisers. Each EU entry names one strain by its deposit number, its holder, the animal categories it covers and a legal minimum content in colony-forming units (CFU) per kg of complete feed at 12 % moisture, or per litre of drinking water.

The organisms fall into three groups: spore formers such as Bacillus and Clostridium butyricum, lactic acid bacteria such as Enterococcus and Pediococcus, and live yeast (Saccharomyces cerevisiae). In the US no direct-fed microbial holds an FDA approval; a product stays food under Compliance Policy Guide Sec. 689.100 only while its species is AAFCO-listed and its label makes no drug claim. Only one EU strain, 4b1841, carries a legal maximum, and only one, the live yeast 4d1703, carries a Salmonella-reduction claim.

Every value on this page is compiled from the EU authorising acts, the EU Register of Feed Additives, EFSA opinions, the FDA compliance policy guide and named peer-reviewed meta-analyses, checked on 3 November 2026; see the methodology.

What Are Direct-Fed Microbials?#

A direct-fed microbial is a feed additive that contains a declared number of viable microorganisms of a named strain, fed so that the organisms act in the gut or rumen of the animal. Regulation (EC) No 1831/2003 names "micro-organisms" in its feed additive definition (Article 2(2)(a)), and live microbials form a zootechnical group among the feed additive categories of that regulation. Synonyms include DFM, gut flora stabilisers and, in Canada, viable microbial products (VMP); "eubiotics" is a marketing umbrella, not a legal term.

EU functional group 4b and the US direct-fed microbial classification#

In the EU, probiotics belong to category 4 zootechnical additives, mostly to functional group 4b gut flora stabilisers, defined as "micro-organisms or other chemically defined substances, which, when fed to animals, have a positive effect on the gut flora". "Probiotic" is not a legal category; the five functional groups of category 4 are listed below.

• 4a digestibility enhancers (for example live yeast CBS 493.94 as 4a1704)
• 4b gut flora stabilisers (most bacterial and yeast strains)
• 4c substances that favourably affect the environment
• 4d other zootechnical additives (for example the Salmonella claim of 4d1703)
• 4e physiological condition stabilisers (Bifidobacterium longum CNCM I-5642 for dogs, 4e001)

In the US, direct-fed microbials have no approval route of their own. FDA's Compliance Policy Guide Sec. 689.100, issued on 2 May 1988 and revised in March 1995, treats products "purported to contain live (viable) microorganisms" as food when the species is AAFCO-listed and no drug claim is made.

Category 4 authorisations are holder-bound, so each EU probiotic entry carries a company name, such as Toa Biopharma Co., Ltd. for 4b1895.

Why live microorganisms are added to feed#

Feed mills add live microorganisms to stabilise the gut or rumen microbiota, a function the EU accepts only after EFSA FEEDAP has assessed efficacy for each target category. EFSA assesses each strain under its characterisation guidance (EFSA Journal 2018;16(3):5206) and efficacy guidance (EFSA Journal 2024;22(7):8856).

EU feed additive labels may not carry disease claims (Article 13(3) of Regulation (EC) No 1831/2003), and no EU gut flora stabiliser authorisation carries a necrotic-enteritis claim. Which additive acts in which gut section is set out under gut health in livestock and poultry.

What is not a probiotic in feed law#

A product without a viable-count guarantee is not a probiotic in feed law: MOS, yeast cell walls, yeast beta-glucans and inactivated bacteria are EU feed materials, and postbiotics are by definition inanimate. The EU Catalogue of feed materials (Regulation (EU) 2022/1104) lists them, and no feed additive authorisation for them was found as of 3 November 2026. The ISAPP consensus (Salminen and colleagues, 2021) defines a postbiotic as "a preparation of inanimate microorganisms and/or their components that confers a health benefit on the host".

Technological microorganisms are not probiotics either: Lactiplantibacillus plantarum DSM 11520 is the acidity regulator 1j002 (to 2034-05-08), a Bacillus subtilis and Lactococcus lactis preparation is the hygiene condition enhancer 1n01 (to 2034-03-21), and silage inoculants sit in group 1k. The Aspergillus oryzae NRRL 458 fermentation product 4a2i for dairy cows (min 85, max 300 mg/kg, Implementing Regulation (EU) 2023/1711, to 2033-09-28) is a digestibility enhancer, not a probiotic.

In the US, 21 CFR 573.587 lists dried Komagataella pastoris yeast as a non-viable protein source for broiler chickens at no more than 10 % of the formulation (89 FR 72315, 5 September 2024), so it is not a direct-fed microbial. Inactivated yeast and cell-wall products are covered under prebiotics, yeast derivatives and postbiotics.

Probiotic Member Table: Organism Groups, Strain Sections and Species Pages#

The six additive rows below cover the three organism groups, the two Clostridium butyricum preparations described on this page and the enzyme-Bacillus combination, with EU entry, categories, legal minimum and US status side by side; the three species pages follow in the second table.

Contents in the first table are legal minimum contents per kg complete feed (12 % moisture), not recommended doses; no maximum content is set except for 4b1841 in pigs, and no withdrawal period is set. The species pages carry the strain tables by animal category.

How Do Direct-Fed Microbials Work?#

Direct-fed microbials work only while alive: spores germinate in the small intestine, vegetative bacteria colonise and acidify the gut, and yeasts act in the rumen or hindgut.

Gut colonisation, competitive exclusion and rumen fermentation#

Spore-forming strains survive pelleting and gastric passage, germinate in the small intestine and secrete enzymes and antimicrobial lipopeptides, while lactic acid producers lower gut pH and compete for adhesion sites. At class level, spore formers competitively exclude Clostridium perfringens and coliforms, although claims vary by strain, and lactic acid bacteria add bacteriocins. Clostridium butyricum is a butyrate-producing spore former.

Live yeast scavenges oxygen in the rumen and stimulates fibrolytic and lactate-utilising bacteria, with higher rumen pH and volatile fatty acids. In horses the target is the hindgut: in one trial, live yeast CBS 493.94 increased caecal fibrolytic enzyme activities of solid-adherent bacteria (Jouany and colleagues, 2009, Journal of Animal Science).

Why the strain, not the species, decides the effect#

An EU authorisation covers one strain, identified by species and culture-collection deposit number, so two Bacillus subtilis products with different deposit numbers are two different additives, and legal minima between strains differ about 100-fold. Four EU acts have renamed authorised strains, and labels may lag behind them:

• Bacillus subtilis C-3102 became Bacillus velezensis DSM 15544 (Implementing Regulation (EU) 2022/703);
• Bacillus licheniformis DSM 5749 became Bacillus paralicheniformis DSM 5749 (2023/2647);
• Bacillus coagulans DSM 32016 became Weizmannia faecalis DSM 32016 (2024/1054);
• Enterococcus faecium NCIMB 10415 became Enterococcus lactis NCIMB 10415 (2024/1161).

Safety status differs by species too: on the EFSA qualified presumption of safety (QPS) list of 8 July 2025, Enterococcus faecium and Clostridium butyricum are not recommended for QPS, while Bacillus subtilis, B. licheniformis, B. velezensis, B. amyloliquefaciens, B. pumilus, S. cerevisiae and the listed lactic acid bacteria are QPS with qualifications.

Evidence from meta-analyses#

Probiotics raised broiler body weight by 66.6 g and cut feed conversion ratio by 0.058 across 106 trials in the meta-analysis by Bilal and colleagues (Poultry Science, 2026), with Bacillus strains more consistent than Lactobacillus (feed conversion improved at days 21, 35 and 42 versus day 21 only). In piglets, Lactobacillus-based probiotics improved feed intake, daily gain, gain-to-feed and villus height over 20 studies (Zhu and colleagues, 2022, Frontiers in Veterinary Science). In growing-finishing pigs, Bacillus spp. reduced feed-to-gain (SMD -0.127, p < 0.001) without a significant effect on intake or gain (Gonzalez-Ronquillo and colleagues, 2022, Animals).

Across 157 ruminant experiments, live yeast raised rumen pH by 0.03, volatile fatty acids by 2.17 mM, organic matter digestibility by 0.8 %, dry matter intake by 0.44 g/kg body weight and milk by 1.2 g/kg body weight (Desnoyers and colleagues, 2009, Journal of Dairy Science). In beef cattle, DFM cut the odds of faecal E. coli O157 shedding (OR 0.46, 95 % CI 0.36 to 0.60; Wisener and colleagues, 2015, Zoonoses and Public Health), and in calves, lactic acid bacteria cut diarrhoea incidence (RR 0.437, 95 % CI 0.251 to 0.761; Signorini and colleagues, 2012, Research in Veterinary Science). These responses are weighed against the other alternatives to antibiotic growth promoters.

Two caveats limit these numbers. Blajman and colleagues (2014, British Poultry Science) found a body-weight-gain effect of SMD 0.661 over 48 trials that was no longer significant (0.059) after trim-and-fill correction for publication bias, although water delivery outperformed feed. Yeast culture results, such as 1.18 kg/d more milk in Poppy and colleagues (2012, Journal of Dairy Science), do not transfer to live yeast. All of these are study results, not legal claims.

Which Probiotic for Which Feed Process and Claim?#

The feed process picks the organism group before the species does: spores for pelleted and extruded feed, coated lactic acid bacteria and yeast or drinking-water entries where heat would kill vegetative cells. The claim narrows the choice further, because only the act decides what a strain may be sold for.

Selection follows a fixed order: the act names the exact animal category (EU) or the species is AAFCO-listed without a drug claim (US), the form matches the process, the coccidiostat and acid compatibility list fits, and the final feed stays at or above the legal minimum after process losses. Prebiotics feed the resident flora and postbiotics carry no live count, differences set out attribute by attribute in probiotics vs prebiotics vs postbiotics in animal feed.

Selection by Species and Diet#

Each EU entry names its target categories, so a strain authorised for chickens for fattening may not be fed to laying hens or dairy cows unless its act names them. Fish and crustaceans have one entry, 4d1712, and calf probiotics are covered with milk replacer and starter feed on the calf page. The table lists the entries per species.

Probiotics for cattle and dairy cows#

Dairy cows have EU entries in two organism groups: live Saccharomyces cerevisiae strains with legal minima from 1 × 10^7 CFU/kg (4a1704) to 2 × 10^9 CFU/kg (4b1710), and the Bacillus entry 4b1896 at 3.8 × 10^8 CFU/kg complete feed. 4b1896, a Bacillus paralicheniformis and B. subtilis preparation held by Chr. Hansen A/S under Implementing Regulation (EU) 2025/2511, is the first EU Bacillus gut flora stabiliser for dairy ruminants and can also be given via drinking water. NCYC R404 (4b1871, Volac International Ltd) for cows runs to 2036-02-15 (2026/155).

No single strain is the best probiotic for cattle in EU law; the right one is the entry whose act names the animal category. Cattle for fattening have three yeast entries (4b1702, 4b1710, 4b1711) and calves for rearing two (4b1702, 4b1711), with the minima in the table. The Desnoyers and Wisener meta-analyses above cover live yeast and DFM responses in cattle.

Horses, sheep, goats and rabbits#

Live yeast strains carry EU entries for horses with legal minima from 8 × 10^8 CFU/kg (4b1702) to 3 × 10^9 CFU/kg complete feed (4b1711, 4b1710, 4d24), and CBS 493.94 (4a1704, 1.6 × 10^9 CFU/kg) sits in functional group 4a digestibility enhancers.

Small ruminants and rabbits rely on the same yeasts: 4b1711 names dairy goats and dairy sheep (Implementing Regulation (EU) 2019/857, to 2029-06-17), 4b1702 names dairy goats, dairy sheep and lambs for fattening (2019/899, to 2029-06-23), and 4b1702 also covers rabbits other than suckling and breeding rabbits (2025/275, to 2035-03-05).

Dogs and cats#

Pet food manufacturers can use strains with EU entries for dogs and cats, including the highest legal minimum in the group, 7 × 10^10 CFU/kg for S. cerevisiae MUCL 39885 (4b1710) in dog feed (2022/272) and cat feed (2024/980). Bifidobacterium longum CNCM I-5642 (4e001) is the one physiological condition stabiliser among them, authorised for dogs at 3.5 × 10^9 CFU/kg by Implementing Regulation (EU) 2025/1776 to 2035-09-29, with Nestlé Enterprises S.A. - Division Nestlé Purina Petcare Europe as holder.

Dosing Logic: Legal Minimum CFU, the One Legal Maximum and Trial Doses#

EU probiotic entries set a legal minimum content in CFU per kg of complete feed at 12 % moisture, from 1 × 10^7 to 7 × 10^10 CFU/kg depending on strain and species, and only one strain carries a legal maximum. That strain is Enterococcus lactis DSM 7134 (4b1841) in pigs under Implementing Regulation (EU) 2020/159, to 2030-02-26. The table gives each figure with its basis.

A legal minimum is the lowest content at which the authorisation applies, not a recommended dose or an optimum, and per-animal-per-day values cannot be compared with CFU/kg values without the feed intake. The 4b1896 per-day figure applies only via drinking water, and the median 486 mg/d of Bacillus spp. in the Gonzalez-Ronquillo pig meta-analysis describes the trials, not a CFU content.

Regulation Summary: EU Authorisations and US Status#

An EU probiotic needs a strain-specific, holder-bound authorisation under Regulation (EC) No 1831/2003, while in the US no direct-fed microbial holds an FDA approval and the product stays food only without drug or structure/function claims.

EU entries, holders and renewal status#

Every EU probiotic entry names its holder, target categories, legal minimum and end date, and the entries 4b1707, 4b1715 (laying hens), 4b1820 (ornamental fish) and 4d12 have printed expiries before 3 November 2026. For each, the EU Register records a renewal application and still lists the entry pending the Commission decision (Article 14 of Regulation (EC) No 1831/2003).

Two further entries have renewal applications noted before their printed expiries: Enterococcus faecium CECT 4515 (4b1713, Implementing Regulation (EU) 2017/961, printed expiry 2027-06-28) and the B. subtilis DSM 5750 and B. paralicheniformis DSM 5749 preparation (4b1700i, 2017/447, printed expiry 2027-04-04). Article 14 renewals are explained under EU feed additive regulation.

US status: direct-fed microbials under CPG Sec. 689.100#

In the US, a direct-fed microbial is food under section 201(f)(3) of the FFDCA when its species is on the AAFCO Official Publication microorganism list and its label makes no disease or structure/function claim. CPG Sec. 689.100 sorts every product into one of three legal classes.

• Food: species AAFCO-listed, no drug or structure/function claim.
• New animal drug: any disease or structure/function claim, adulterated under section 501(a)(5) unless an approved NADA covers it.
• Food additive: an organism not on the list, adulterated under section 402(a)(2)(C) unless a food additive regulation exists.

The label carries the statement "Contains a source of live (viable) naturally occurring microorganisms", followed by each organism and its guarantee in CFU/g (Regulation 4(g) of the Uniform State Feed Bill). US listing works at species level, so the strain is not required on the label, unlike an EU entry tied to a deposit number.

The species list itself sits in the AAFCO Official Publication, and EU authorisation of a strain does not imply US status, nor the reverse.

Great Britain and Canada#

Great Britain retained the EU authorisations in force at 31 December 2020, and later EU authorisations do not apply there automatically. GB entries are listed in the FSA register of regulated feed additive products. In Canada, each viable microbial strain needs CFIA pre-market registration (RG-1 section 3.22), and production claims beyond NRC normal ranges, any reference to intestinal flora or a disease claim make the viable microbial product a veterinary drug.

Sourcing Checklist for Direct-Fed Microbials#

Source a direct-fed microbial in seven checks, from the deposit number on the label to the CFU count at the end of shelf life.

1. Match the identification number, strain and deposit number to the act, allowing for renamed species.
2. Confirm that the act names the target category, for example "chickens for fattening" rather than "poultry".
3. Ask for the CFU/g guarantee at the end of shelf life and compare it with the additive minimum in the act (at least 5 × 10^8 CFU/g for 4b1830).
4. Request pelleting, premix and storage stability data; for one Bacillus preparation EFSA reports under 0.5 log loss after 6 months in premix at 25 °C and 60 % relative humidity.
5. Check the coccidiostat compatibility list in the act; for 4b1822 in chickens it names only diclazuril, monensin sodium and nicarbazin.
6. Ask for the enumeration method on the certificate of analysis: EN 15784 Bacillus, EN 15788 Enterococcus, EN 15787 Lactobacillus and Weizmannia faecalis, EN 15786 Pediococcus, EN 15785 Bifidobacterium, EN 15789 S. cerevisiae, ISO 15213 C. butyricum; CEN/TS 17697 or sequencing (CEN/TS 15790 for yeast) for identity.
7. Ask for FAMI-QS or GMP+ certification and the user-safety data.

Offers become comparable when they name the identification number, the guaranteed CFU/g at expiry and the form, such as a spore powder or coated live yeast.

Direct-Fed Microbial Questions#

Three questions about direct-fed microbials come up whenever a probiotic enters a feed programme.

What should you not mix with probiotics?#

Do not combine a probiotic with a coccidiostat its act does not list, with high-dose antibiotics or with a conditioning temperature that kills vegetative cells. Vegetative lactic acid bacteria and uncoated yeast added before an 85 to 90 °C conditioner do not keep their label CFU in the final feed.

Each act lists its compatible coccidiostats in "other provisions", so the list differs from strain to strain. Approved uses, programmes and withdrawal for coccidiostats sit on their own page.

Is there a list of direct-fed microbials?#

Yes, two: the EU Register of Feed Additives lists each authorised strain with its identification number, and in the US the AAFCO Official Publication lists the microorganism species accepted for direct-fed microbials. The EU Register entry names the holder, categories, legal minimum and act for each strain, while the AAFCO list works at species level and names no holder.

Do probiotics for animals have side effects?#

EFSA concludes on target-animal safety strain by strain before an EU entry is granted, and the acts treat microbial preparations as potential skin and respiratory sensitisers for the people who handle them. The acts require operational procedures or breathing and skin protection for users. Microorganisms carry no harmonised CLP classification, so hazard details come from the supplier SDS, and no maximum residue limit is set for consumers.