Perspectives and advances in probiotics and the gut microbiome in companion animals
REVIEW
J Anim Sci Technol 2022;64(2):197-217
Journal of Animal Science and Technology
https://doi.org/10.5187/jast.2022.e8 pISSN 2672-0191 eISSN 2055-0391
Daniel Lee1#, Tae Wook Goh1#, Min Geun Kang1, Hye Jin Choi1
,
So Young Yeo1, Jungwoo Yang2, Chul Sung Huh3,4, Yoo Yong Kim1 and
Younghoon Kim1*
1Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Science, Seoul
National University, Seoul 08826, Korea
2Ildong Bioscience, Pyeongtaek 17957, Korea
3Research Institute of Eco-Friendly Livestock Science, Institute of Green-Bio Science and Technology,
Seoul National University, Pyeongchang 25354, Korea
4Graduate School of International Agricultural Technology, Seoul National University, Pyeongchang
25354, Korea
Received: Nov 23, 2021
Revised: Jan 14, 2022
Accepted: Jan 17, 2022
# These authors contributed equally to
this work.
*Corresponding author
Younghoon Kim
Department of Agricultural
Biotechnology and Research Institute
of Agriculture and Life Science, Seoul
National University, Seoul 08826,
Korea.
Tel: +82-2-880-4808
E-mail: ykeys2584@snu.ac.kr
Copyright © 2022 Korean Society of
Animal Sciences and Technology.
This is an Open Access article
distributed under the terms of the
Creative Commons Attribution
Non-Commercial License (http://
creativecommons.org/licenses/by-
nc/4.0/) which permits unrestricted
non-commercial use, distribution, and
reproduction in any medium, provided
the original work is properly cited.
ORCID
Daniel Lee
https://orcid.org/0000-0001-7224-3958
Tae Wook Goh
https://orcid.org/0000-0003-4705-3348
Min Geun Kang
https://orcid.org/0000-0002-2204-6443
Hye Jin Choi
https://orcid.org/0000-0002-5977-2780
Abstract
As the number of households that raise dogs and cats is increasing, there is growing interest
in animal health. The gut plays an important role in animal health. In particular, the microbi-
ome in the gut is known to affect both the absorption and metabolism of nutrients and the
protective functions of the host. Using probiotics on pets has beneficial effects, such as mod-
ulating the immune system, helping to reduce stress, protecting against pathogenic bacteria
and developing growth performance. The goals of this review are to summarize the relation-
ship between probiotics/the gut microbiome and animal health, to feature technology used
for identifying the diversity of microbiota composition of canine and feline microbiota, and to
discuss recent reports on probiotics in canines and felines and the safety issues associated
with probiotics and the gut microbiome in companion animals.
Keywords: Probiotics, Gut microbiome, Companion animal, Canine, Feline
INTRODUCTION
Terms such as ‘companion animals’ apply to households with pets, companion dogs and companion
cats that are frequently encountered in the surroundings. The word ‘companion’, with which we are
already familiar, refers to an animal that lives with humans and was first proposed by zoologist and
Nobel Prize winner Konrad Lorenz at an international symposium held in Vienna, Austria in 1983
[1]. Households that raise these pets accounted for 29.7% of the total households in Korea, with 6.04
million households at the end of 2020 [2]. As the number of people raising companion animals is
increasing, the relationship between humans and companion animals is further developing.
Most pet owners currently treat their pets as family, colleagues, and friends [3]. Pet humanization,
a phenomenon that recognizes companion animals as family members and treats them as individuals
with emotions, has been established as a global trend [4]. In Korea, the trend of pet humanization is
https://www.ejast.org
197Probiotics and the gut microbiome in companion animals
So Young Yeo
https://orcid.org/0000-0002-8448-1408
Jungwoo Yang
https://orcid.org/0000-0003-3836-729X
Chul Sung Huh
https://orcid.org/0000-0003-0287-2411
Yoo Yong Kim
https://orcid.org/0000-0001-8121-3291
Younghoon Kim
https://orcid.org/0000-0001-6769-0657
Competing interests
No potential conflict of interest relevant to
this article was reported.
Funding sources
This work was supported by the National
Research Foundation of Korea (NRF) Grant
funded by the Korean government (NRF-
2021R1A2C3011051).
Acknowledgements
Not applicable.
Availability of data and material
Upon reasonable request, the datasets
of this study can be available from the
corresponding author.
Authors’ contributions
Conceptualization: Lee D, Goh TW, Kang
MG, Choi HJ, Yeo SY, Yang J, Huh CS,
Kim YY, Kim Y.
Data curation: Lee D, Goh TW, Kim Y.
Formal analysis: Lee D, Goh TW, Kim Y.
Writing - original draft: Lee D, Goh TW, Kang
MG, Choi HJ, Yeo SY, Yang J, Huh CS,
Kim YY, Kim Y.
Writing - review & editing: Lee D, Goh TW,
Kang MG, Choi HJ, Yeo SY, Yang J, Huh
CS, Kim YY, Kim Y.
Ethics approval and consent to participate
This manuscript does not require IRB/IACUC
approval because there are no human and
animal participants.
also spreading, with 88.9% of companion households and 64.3% of general households agreeing
to the phrase ‘pets are part of the family’ [2]. A typical example is that the pets do the same things
as humans do, such as having birthday parties for dogs and cats, sleeping with the owner in the bed,
and others. Companion animals have become an increasingly important part of human life, and
therefore, the health and well-being of pets have increasingly attracted interest in recent decades [5].
Dogs and cats have evolved into carnivores with high-protein diets and have relatively simple
gastrointestinal tracts (GITs) [5,6]. Cats are carnivores that rely on high-protein animal tissues to
meet their unique nutritional requirements in the wild and consume protein-containing feed to
meet their nutrients in the case of household felines. They are metabolically adapted to low glucose
utilization and high protein metabolism [5,7]. Although dogs share many anatomical and metabolic
characteristics with cats, they are metabolically omnivorous and can digest, absorb and metabolize
significant amounts of carbohydrates [8].
The gut plays an important role in animal health, and the GIT contains a complex microbial
community. A healthy gut is known to affect host physiology and well-being. This microbial
ecosystem acts in several ways, affecting both the absorption and metabolism of nutrients and the
protective functions of the host. Probiotics are defined as ‘living microorganisms that provide health
benefits to the host when administered in appropriate amounts’ [9]. Recently, gut-related probiotic
products aimed at pets, particularly dogs and cats, have also gained in popularity among owners
[10]. The benefits of using probiotics for pets include their modulation of the immune system,
help in reducing stress, protection against infections caused by intestinal pathogens and growth
performance development [11]. As dogs and cats become family members, the number of studies
about dogs and cats has been increasing. Among their topics, knowledge about the gut microbiome
and probiotics in dogs and cats is still expanding. However, published papers on the application
of probiotics in companion animals are significantly limited compared to those in humans. The
purpose of this review is to describe the current knowledge about the gut microbial communities in
dogs and cats in relation to probiotics.
PROBIOTICS FOR COMPANION ANIMALS
Probiotics that are living and beneficial microbiota have been used for companion animal’s health
[9]. As people’s desire to have their pets for a long time has increased, interest in probiotics has
also attracted more attention [10]. Probiotics provide beneficial health effects to the host animal
by altering the gastrointestinal (GI) flora. The GI benefits for dogs and cats include maintaining a
balanced and healthy gut microbiome, preventing diarrhea, and managing small intestinal bacterial
overgrowth and inflammatory bowel disorders [12]. Since dogs and cats have different dietary
needs and digestive systems, their needs for and effects from probiotics differ.
Probiotics for canines
Canines are considered animal models for the study of the human microbiome because of the high
structural and functional similarity between the canine and human microbiomes [13]. The study
of the dog microbiome can be predictive of the human microbiome. Thus, the study of dogs offers
two advantages not only directly for dogs but also for its potentially benefits for humans [14].
Although the beneficial effects of probiotics have been extensively studied in humans and animals,
the exact mechanisms of probiotic-based immune modulation are not entirely clear, and the efficacy
of probiotic applications varies depending on many different factors [15]. Recent reports of using
probiotics in canines are shown in Tables 1 and 2.
GI disorders are one of the most common health problems in dogs [16,17]. Regardless of the
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cause, most GI disorders present with acute or chronic diarrhea, or, in some cases, vomiting or
anorexia [5,18,19]. Many previous studies have shown positive results regarding the treatment of
dogs with different types of probiotics [20,21]. Dogs on diets supplemented with 2 × 1010 CFU/
day canine-derived probiotic Bifidobacterium animalis AHC7 had a significantly more rapid
resolution of acute diarrhea than dogs that received placebo [22]. The administration of Lactobacillus
rhamnosus MP01 and L. plantarum MP02, two strains isolated from canine milk, decreased the
Faecalibacterium in feces [23]. Supplementing 5 × 109 CFU/day of L. murinus LbP2 in dogs
improved their stool output, fecal consistency, mental status, and appetite compared to the control
[24]. A total of 15 adult female dogs who were given 2.3 × 108 CFU/day canine-origin probiotic
L. johnsonii CPN23 exhibited increased fiber digestibility and concentrations of short-chain fatty
acids in their feces and reduced fecal ammonia concentrations compared to the control [25]. Dogs
that consumed 107–109 CFU/day canine-derived probiotic L. fermentum CCM 7421 displayed an
increased lactic acid bacteria population, reduced Clostridia population and some gram-negative
bacterial genera. Additionally, dogs that consumed probiotics showed improved total protein,
cholesterol and alanine transaminase in blood samples [26]. Three milliliters of 109 CFU/mL of
the new potential probiotic L. fermentum AD1 significantly increased total lipids and total protein
and significantly decreased the glucose concentration in the bloodstream [27]. Dogs fed 1.04 ×
109 CFU/mL B. animalis B/12 showed a significantly decreased concentration of triglycerides and
albumin and increased acetic, acetoacetic, and valeric acid in feces [28]. Supplementing 108 CFU/
mL canine-origin probiotic L. johnsonii CPN23 in adult female dogs decreased their plasma glucose
and cholesterol levels and increased the high-density lipoprotein and low-density lipoprotein ratio
[29]. Dogs receiving Enterococcus faecium DSM 32820 had optimal fecal consistency throughout the
experiment, significantly stimulated phagocytic activity and a metabolic burst activity of leukocytes
and lower serum glucose concentrations [30]. Heathy dogs receiving 5 × 109 CFU/kg L. acidophilus
D2/CSL showed higher body condition scores than the control dogs and there was a positive effect
on their fecal consistency [31]. The probiotic feed additive contained three different bacterial strains,
namely, L. casei Zhang, L. plantarum P-8, and B. animalis subsp. lactis V9 promoted the average
daily feed intake, improved average daily weight gain, increased beneficial bacteria and decreased
potentially harmful bacteria [14]. The probiotic E. faecium SF68 improved diarrhea symptoms
compared to the control, and Giardia cysts were eliminated [32]. Adding 5 × 108 CFU/day E.
faecium SF68 significantly increased the triglyceride concentration and decreased the cholesterol
concentration [33].
The gut microbiome greatly affects the health and disease of the host so maintaining it in good
condition is important for the health of the host [34]. Many factors influence the composition of
the gut microbiome and aging is one of the greatest impacts [35]. After all, this aging which is
defined as the gradual changes that occur after maturation in various organs, resulting in decreased
functional capacity in the gut microbiome is thought to be somehow related to the health of the
host [36]. Masuoka et al. [34] conducted the experiment with dogs of 5 different age groups (pre-
weanling, weanling, young, aged and senile) and analyzed the composition of their intestinal
microbiota of dogs in different age groups. As a result, the composition of the dog’s intestinal
microbiota changed with age. Lactobacillus and Bifidobacterium were found to decrease as the dog
aged. This experiment showed that the gut microbiome of dogs can be changed regarding the age at
the level of bacterial groups and species. Further studies are needed to be done to identify whether
different probiotics are needed for different phases of life.
https://doi.org/10.5187/jast.2022.e8 https://www.ejast.org | 199Probiotics and the gut microbiome in companion animals
Probiotics for felines
Cats have trillions of live bacteria in their bodies, which are mostly in their intestines [21]. Each
cat’s bacterial population is different for individuals and can be changed based on diet, health
status, and lifestyle choices [37,38]. During times of stress and infection, the microbiome balance
can increase the number of bad bacteria, disrupting the system’s balance and potentially causing
digestive problems such as decreased appetite, vomiting, diarrhea or stool changes [39,40].
Supplementing probiotics for felines can be one of the best ways to add good bacteria to the cat
body [21]. Recent reports of using probiotics in felines are listed in Tables 1 and 2.
Although many studies have investigated the use of probiotics in dogs, studies in cats are
relatively scarce. Few studies on probiotic usage in cats have been reported to date, and because of
differences in host physiologic characteristics and the diet, the probiotic efficacy in cats cannot be
extrapolated from studies in dogs [41]. The purpose of this review paper is to discuss various results
about treating cats with different types of probiotics. Kittens receiving 2.85–4.28 × 108 CFU/day E.
hirae showed high intestinal colonization and fecal shedding of live E. hirae during administration
[42]. Supplementing 2 × 108 CFU/day L. acidophilus DSM13241 as a probiotic in healthy adult
cats increased the numbers of beneficial L. and L. acidophilus groups in feces and decreased the
numbers of Clostridium spp. and E. faecalis. It also decreased the fecal pH and plasma endotoxin
concentrations and resulted in systemic and immunomodulatory changes in treated cats [41].
Kittens fed 5 × 109 CFU/day E. faecium SF68 showed a significantly higher percentage of CD4+
lymphocytes than controls [43]. Healthy adult cats fed 5 × 109 CFU/kg L. acidophilus D2/CSL had
better results in terms of their fecal quality parameters and had increased Lactobacillus counts and
decreased total coliform bacteria counts [44]. The percentage of cats with diarrhea was significantly
lower in the 2.1 × 109 CFU/day E. faecium SF68 group than in the control group [45]. Young
adult cats receiving E. faecium SF68 had significantly lower total diarrhea scores for days 1–11
compared to the control. Additionally, feeding E. faecium SF68 could lessen some associated clinical
abnormalities [46]. Feeding Enterococcus faecium SF68 in cats with chronic feline herpesvirus-1
(FHV-1) infection showed fecal microbial diversity throughout the study which indicates a more
stable microbiome. It also lessened the morbidity associated with chronic FHV-1 infections [47].
Healthy cats with 5 × 109 CFU from a mixture of seven bacterial species per day (Proviable®-DC)
showed an increased abundance of probiotic bacteria in the feces. Probiotics also improved diarrhea
after 21 days of feeding [48]. Cats with chronic gingivostomatitis that were fed 1 × 108 CFU/mL L.
plantarum showed many positive results in gingivostomatitis symptoms. There was an improvement
in the time of recurrence, and the symptoms of chronic feline gingivostomatitis disappeared after
two weeks of administration. Additionally, ulceration, inflammation and oral cavity pain decreased,
and thalism and halitosis disappeared [49]. Giving multistrain probiotic products to 8-month-
old male cats with feline idiopathic cystitis effectively managed this disease due to the effects of
bactericidal, anti-inflammatory, and immunomodulatory actions [50].
The health and disease of the host are affected by gut microbiota, maintaining the gut microbiota
is getting more important as cats get aged. Masuoka et al. [51] conducted the experiment with
cats of 5 different age groups (pre-weanling, weanling, young, aged and senile) and analyzed the
composition of their intestinal microbiota of cats in different age groups. The results suggested that
the composition of the cat’s gut microbiome changed with age, whereas the change was different
from that of dogs. Bifidobacterium which predominated in the gut of dogs did not appear to be
important in the gut of cats. Instead, enterococci appeared to be the main lactic acid-producing
bacteria in cats. Ultimately, the results of this study indicated that the compositions of the gut
microbiome between dogs and cats are different and those compositions are changing with aging.
Not only are different probiotics might need for dogs and cats but also for regarding aging. Further
200 | https://www.ejast.org https://doi.org/10.5187/jast.2022.e8https://doi.org/10.5187/jast.2022.e8 https://www.ejast.org | 201
Table 1. List of canine and feline origin bacterial strains used as probiotic properties in canines and felines
Bacterial strains Amount Source Group Tested for Result Reference
Bifidobacterium animalis AHC7 2 × 1010 CFU/day Canine Young adult dogs with acute
diarrhea
Lactobacillus rhamnosus MP01
109 CFU/day Canine 1 month old puppies Lactobacillus plantarum MP02
Assessment for managing
acute diarrhea
Assessments for preventing
gastrointestinal infection in
puppies
Lactobacillus murinus LbP2 5 × 109 CFU/day Canine Dogs with canine distemper virus
Assessment of fecal and
(CDV)-associated diarrhea
mental status
Lactobacillus johnsonii CPN23 2.3 × 108 CFU/day Canine Adult female Labrador dogs Assessment of nutrient digest-
ibility and fecal fermentative
metabolites
Lactobacillus fermentum CCM
7421
107–109 CFU/day Canine Dogs suffering from gastrointesti-
nal disorder
Assessment of blood samples
and composition of the
fecal microbiome
Lactobacillus fermentum AD1 3 mL of 109 CFU/mL Canine Healthy dogs Assessment of blood sample
and composition of fecal
microbiome
Bifidobacterium animalis B/12 Lactobacillus johnsonii CPN23 Enterococcus faecium DSM
32820
Lactobacillus acidophilus
DSM13241
1 mL of 1.04 × 109
CFU/mL
Canine Healthy dogs Assessment of blood samples
and composition of fecal
microbiome
108 CFU/mL (0.1 mL/
kg BW)
Canine Adult female dogs Assessment of blood sample
profile
109 CFU/day Canine Healthy dog Assessment of blood sample
profile
2 × 108 CFU/day Feline Healthy adult cats Assessment for improving
intestinal health in cats
Enterococcus hirae 2.85–4.28 × 108 CFU/
day
Feline Kittens Assessment for preventing
atypical Enteropathogenic
E. coli (EPEC) in kittens
Enterococcus faecium SF68 5 × 109 CFU/day Feline Kittens Effects of Enterococcus
faecium strain SF68 sup-
plementation on immune
function
Reduced diarrhea compared to placebo group [22]
Significantly increased Lactobacillus and Faecalibac-
terium in fecal
Significantly increased SCFAs concentration in feces
Prevented gastrointestinal infection
Fecal consistency, mental status and appetite were
significantly improved
Increased crude fiber digestibility
Increased concentrations of SCFAs in feces
Reduction in fecal ammonia concentration
[23]
[24]
[25]
Improved total protein, cholesterol and ALT in blood
samples
Increased lactic acid bacteria population and de-
creased clostridia population along with some of
the gram-negative bacterial genera
Modulate liquid feces to normal consistency (dogs
with diarrhea)
Significantly increased total lipid and total protein in
the blood
Significantly decreased glucose concentration in the
bloodstream
Significantly increased the number of lactobacilli and
enterococci in the feces
[26]
[27]
Significantly decreased concentration of triglycerides
and albumin in blood serum
Increased ALT and ALP
Increased acetic, acetoacetic and valeric acids in
feces
[28]
Decreased plasma glucose and cholesterol level
Increased HDL/LDL ratio
[29]
Decreased serum glucose concentration [30]
Increased numbers of beneficial Lactobacillus and L.
acidophilus groups in feces and decreased num-
bers of Clostridium spp. and Enterococcus faecalis
Decreased fecal pH and plasma endotoxin concen-
trations resulting in systemic and immunomodula-
tory changes in treated cats
Highly effective at promoting intestinal colonization
and fecal shedding of live E. hirae during adminis-
tration.
Ameliorated the effects of atypical EPEC experimen-
tal infection on intestinal function and water loss
[41]
[42]
The percentage of CD4+ lymphocytes was signifi-
cantly higher in the treatment group
[43]
SCFAs, short-chain fatty acids; ALT, alanine aminotransferase; ALP, alkaline phosphatase; BW, body weight; HDL, high-density lipoprotein; LDL, low-density lipoprotein.
Lee et al.202 | https://www.ejast.org https://doi.org/10.5187/jast.2022.e8
Table 2. List of bacterial strains of non-canine and non-feline origin when used for their probiotic properties in canines and felines
Bacterial strains Amount Source Group Tested for Result Reference
Lactobacillus casei Zhang
Lactobacillus plantarum P-8
Bifidobacterium animalis subsp.
Lactis V9
2 × 109 CFU/g (2 g for young, 4 g for
training, 10 g for elderly dogs)
[14]
Lactobacillus acidophilus D2/
CSL
Enterococcus faecium SF68 Enterococcus faecium SF68 Lactobacillus acidophilus D2/
CSL (CECT 4529)
Enterococcus faecium SF68 Enterococcus faecium SF68 Enterococcus faecium SF68 5.0 × 109 CFU/kg of diet 5 × 108 CFU/day 5 × 108 CFU/day 5 × 109 CFU/kg of food 2.1 × 109 CFU/day 1/4 can of canned food mixed with
Enterococcus faecium
5 × 108 CFU/day Lactobacillus casei Zhang
(koumiss)
Lactobacillus plantarum P-8
(Fermented dairy products
in China)
Bifidobacterium animalis subsp.
Lactis V9
(Feces of a healthy Mongolian
child)
Gastrointestinal (GI) tract of a
healthy adult chicken
Feces of a healthy breast-fed
baby
Feces of a healthy breast-fed
baby
Conventional foods such as
milk, yogurt and dietary
supplements
A healthy breast-fed newborn
baby
A healthy breast-fed newborn
baby
A healthy breast-fed newborn
baby
Young, training and
elderly dogs
Assessment of nutrition, immunity and
composition of fecal microbiome
Healthy dogs Assessment of nutritional and fecal
status
Dogs with diarrhea Assessment of the effect of administer-
ing metronidazole with Enterococcus
faecium SF68 to treat diarrhea
Healthy dogs Assessment of blood sample profile Healthy adult cats Assessment of the effects on nutritional
condition and fecal quality
Cats Effects on Enterococcus faecium SF68
in diarrhea
Young adult cats Description of the GI abnormalities
associated with the administration of
amoxicillin-clavulanate to cats and
an assessment of whether feeding
with Enterococcus faecium SF68
could ameliorate those abnormalities
Cats with chronic Feline
Assessment of the effect of feeding En-
Herpes virus-1 (FHV-
terococcus faecium SF68 in clinical
1) infection
signs of FHV-1 infection
Promoted the average daily feed intake of elderly dogs
Improved average daily weight gain om all dogs
Enhanced the level of serum IgG, IFN-α, and fecal secretory IgA
(sIgA), while reducing the TNF-α.
Increased beneficial bacteria and decreased potentially harmful
bacteria
Higher body condition score than control group
Positive effect on fecal consistency
Dual therapy that administrates metronidazole with Enterococcus
faecium SF68 improved diarrhea more than administering
metronidazole alone
Giardia cysts were eliminated from the dual treatment group
Mean cholesterol concentration significantly decreased
Mean triglyceride concentration significantly increased
Improved fecal quality parameters
Increased Lactobacillus count and decreased total coliform bacteria
counts
The percentage of cats with diarrhea was significantly lower in the
probiotic group when compared with the placebo group
The total diarrhea scores for days 1–11 were significantly lower in
the cats fed Enterococcus faecium SF68 compared to the cats
fed the placebo
Feeding Enterococcus faecium SF68 can lessen some associated
clinical abnormalities
Fecal microbial diversity was maintained throughout the study in
cats supplemented with Enterococcus faecium SF68, indicating
a more stable microbiome in cats receiving Enterococcus
faecium SF68
Lessened morbidity associated with chronic FHV-1 infection in
some cats
[31]
[32]
[33]
[44]
[45]
[46]
[47]
Proviable®-DC (7 bacterial
species)
5 × 109 CFU of a mixture of seven
bacterial species per day
Lactobacillus plantarum 1 × 108 CFU/mL Mare’s milk Cats with chronic gingiv-
Multistrain probiotic product Adult cat Improvement in stool character Improved diarrhea symptoms after 21-day feeding [48]
ostomatitis
Assessment of preventive and thera-
peutic oral pathology
The administration of the probiotic to the two immunosuppressed
cats affected by gingivostomatitis led to an improvement in the
time of recurrence
The symptoms of chronic feline gingivostomatitis disappeared after
two weeks of administration
The ulceration, inflammation and pain of the oral cavity decreased,
thalism and halitosis disappeared
[49]
Probiotic combination Multistrain probiotic product 8-month-old male cats Management of feline idiopathic cystitis
Probiotic combination treatment effectively managed this disease
(FIC) using probiotics
due the effect of bactericidal, anti-inflammatory, and immuno-
modulatory actions
[50]
Probiotics and the gut microbiome in companion animals
Proviable®-DC (7 bacterial
species)
Lactobacillus casei 4 × 108 CFU,
Lactobacillus rhamnosus 3 × 108 CFU,
Lactobacillus acidophilus 5 × 107 CFU,
Lactobacillus bulgaricus 1 × 107 CFU,
Bifidobacterium infantis 4 × 107 CFU,
Bifidobacterium breve 5 × 107 CFU,
Streptococcus thermophilus 1 × 108 CFU
5 × 109 CFU of a mixture of seven
bacterial species per day
Multistrain probiotic product Healthy cats Assessment of a multispecies on the
fecal microbiome of healthy cats
Increased abundance of probiotic bacteria in the feces of healthy
cats
[85]
IgG, Immunoglobulin G; IFN, Interferon; TNF, tumor necrosis factor.Lee et al.
studies are needed to use different probiotics for different phases of life.
GUT MICROBIOME FOR COMPANION ANIMALS
Gut microbiome and nutrient metabolism
Microorganisms affect the absorption of nutrients in the host and provide beneficial metabolites in
return for using host nutrients [52]. Each intestine harbors a different unique microbial ecosystem
due to anatomical and physiological differences [53]. Additionally, each animal harbors a different
and unique microbial profile. For example, at the species and strain levels, only a few overlap
between individual animals. However, the bacterial phyla, order and genera are shared by most
mammals [54].
The most predominant bacterial gene category in the canine gut is carbohydrate metabolism,
such as that related to mannose, oligosaccharide and raffinose metabolism. The fermentation of
carbohydrates by colonic organisms such as Bacteroides, Roseburia, Ruminococcus and Lachnospiraceae
results in the synthesis of short-chain fatty acids (SCFAs), such as acetate, propionate and butyrate
which are sources of energy for the host [20]. SCFAs have beneficial effects on host health,
including immunomodulatory effects, anti-diarrheic effects, and a regulatory effect on GI motility.
In the case of felines, which are obligate carnivores, consuming raw meat increased Clostridium and
Eubacterium, which are known to produce SCFAs [55].
The synthesis of vitamin K and several components of vitamin B are important functions of
the intestinal microbiota [56,57]. Vitamin K, which is included in fat-soluble vitamins, plays an
important role in prothrombin coagulation factor activity. Therefore, there is a risk of intestinal
bleeding in cases of vitamin K deficiency [58]. Vitamin B12 (also known as cobalamin) is important
for many aspects of a dog’s health [59]. It is crucial for a healthy nervous system and brain function
as well as for the formation and growth of blood cells [60]. Additionally, it is needed to maintain
healthy digestion [61]. As a result of a metagenome analysis using dog feces, genes affecting
lipoprotein lipase activity in adipocytes were identified in intestinal microbial genes, confirming that
microorganisms are also related to lipid metabolism [62].
Gut microbiome and the immune system
The microbiome plays an important role in the immune system of the intestinal tract. In particular,
early microbial exposure significantly affects gut microbiome formation and immune modulation,
which affects susceptibility to intestinal diseases [63]. When comparing animals born through
vaginal delivery with germ-free animals through cesarean section, the germ-free animals have fewer
and smaller peyer’s patches, mesenteric lymph nodes and CD4+ T cells in the lamina propria of
the gut wall [64]. In germ-free animals, a reduction in B cells, macrophages and neutrophils was
confirmed [65]. Additionally, in germ-free animals, immunoglobulin was found at a level of 2%,
which was significantly lower than that in normal healthy animals [66]. The microbiome also plays
a role as a signal indicating health [65]. This characteristic is expected because animals evolved in
coexistence with symbiotic microorganisms for a very long time [67]. Microorganisms that coexist
with animals communicate directly and effectively with their host’s immune system through
metabolites and nutrients [64,65].
Identifying diversity in the canine and feline microbiomes
The intestine is a major part of the body that influences host health. Numerous microbes form the
complex microbial community in the GIT. Disrupting the gut microbiome may cause dysbiosis and
lead to several diseases and disorders, such as diarrhea, allergies and obesity [21]. GI disease caused
https://doi.org/10.5187/jast.2022.e8 https://www.ejast.org | 203Probiotics and the gut microbiome in companion animals
by the dysbiosis of the gut microbial community is also generally observed in dogs and cats [68,69].
Knowing the diversity and taxonomic bacterial distribution of the gut microbiota of healthy dogs
and cats is important as a baseline in future studies evaluating GI diseases in dogs and cats [70,71].
Previous studies have focused on the cultivation of intestinal content to characterize and identify
the microbiota [72–74]. Most of the bacterial groups cultivated from the canine intestine belonged
to Enterobacteriaceae, Bacteroides, Clostridium, Lactobacillus, and Bifidobacterium spp. [75,76].
However, culturing the bacteria to evaluate the complex diversity of the microbiota had limitations.
Bacterial species that can be cultivated by using bacterial culture techniques are only a small portion
of microbiota composition [77,78], anaerobic bacteria can be easily damaged during sample
handling [79,80], the cost used for culture techniques is expensive [80], a great amount of time is
used for isolation and cultivation [80,81]. A novel molecular method that uses the 16S ribosomal
RNA (rRNA)-enabled the evaluation of the diversity and abundance of bacteria in the sample
without culturing [82,83].
The development of next-generation sequencing technologies has helped to characterize bacterial
communities and to understand interactions between hosts and bacteria. Using next-generation
sequencing, dog and cat organ microbiotas have been described. These microbiota include those of
the GIT [70, 84–86], skin [87], oral cavity [88,89], nasal cavity [90], and vagina [91].
Composition of canine and feline microbiome
All animals, including dogs and cats, harbor numerous microorganisms in the GIT [8]. Dogs and
cats have different microbiota compositions and they also differ in the same species [21]. There
are lots of factors that can affect microbiota compositions such as age [92–94], breed [8,95,96],
diet composition [39,92,94], disease [92,93], environment [92,96,97], food type [93,98] and sex
[99,100].
The gut microbiome which is highly related to a healthy life could be affected by dogs’ breed.
There was a relationship between GI conditions and dog breeds [101,102]. According to You and
Kim’s experiment [103], there was a difference in microbial composition in Poodle and Maltese
groups. Also, phylum Fusobacterium was differed by breeds (Maltese, Poodle, and Miniature
Schnauzer). From these data, they suggested that there might be differences in the gut microbiome
composition depending on the dog breeds. According to Lehtimäki et al.’s experiment [104], living
conditions have a significant impact on the skin microbiome in humans and dogs, but not the
gut microbiome. Dogs living in rural and urban environments participated in the study. The skin
microbiome was more diverse among individuals in rural areas compared to urban areas. This study
showed that the living environment had a much greater effect on the skin microbiome than the
guts of dogs. Experiments on changes in the gut microbiome of cats according to breeds and living
environments have been limited. Further research is needed as with dogs. According to Older et
al.’s [96] experiment, breed and living environment played an important role in shaping the cat skin
microbiome. In particular, it seems that the hair coat and grooming according to the cat breeds have
a great influence on the microbiome of the cat’s skin microbiome.
The bacterial count in the stomach is between 104 and 105 CFU/g [105]. In the duodenum and
jejunum, the bacterial counts are generally low (105 CFU/g) but can reach 109 CFU/mL in some
dogs and cats [106]. The ileum contains an increasing number of diverse microbiota, mostly at 107
CFU/mL. The bacterial counts in the colon are between 109 and 1011 CFU/g [38,73].
The healthy canine stomach has a comparably low number of total bacteria. Most belonged
to Proteobacteria (99.6%), and few belonged to Firmicutes (0.3%). The dominant species are
Helicobacter and Lactobacillus spp. [85]. Using 16S rRNA sequences, four phyla (Firmicutes,
Fusobacteria, Bacteroidetes and Proteobacteria) predominated in the small intestine [70]. The
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duodenum of healthy canines consisted of six phyla. Firmicutes predominated followed by
Proteobacteria, Bacteroidetes, Spirochaetes, Fusobacteria and Actinobacteria [37,107]. Healthy dog
microbiota in the jejunum were evaluated, and the most predominant phylum was Proteobacteria
(46%), followed by Firmicutes (15%), Actinobacteria (11.2%), Spirochaetes (14.2%), Bacteroidetes
(6.2%) and Fusobacteria (5.4%) [84]. The ileum microbiota of healthy dogs predominantly consists
of Fusobacteria, Firmicutes and Bacteroidetes [70].
Lactobacillales was present in all parts of the intestines (22% in the duodenum and 10% in the
jejunum). Enterobacterales were more frequently detected in the small intestine than in the colon.
Clostridiales were highly abundant in the duodenum (40%), jejunum (39%), ileum (25%) and
colon (26%) [70]. Facultative anaerobic Lactobacillus strains predominated in the jejunal microbiota,
and L. acidophilus was the most abundant among them [108]. In the jejunal samples, facultative
anaerobic and anaerobic bacteria were similarly detected, while anaerobic bacteria predominated
in the fecal samples. The number of bacteria in the jejunum was 102 to 106 CFU/g, while the
number in the feces was 108 to 1011 CFU/g. Despite the lower number in the small intestine, some
microbial groups were more prevalent in the small intestine than in feces: staphylococci, 64% versus
36%; non fermentative gram-negative rods, 27% versus 9%; and yeasts, 27% versus 5% [73].
Firmicutes, Bacteroidetes, Fusobacteria, Proteobacteria and Actinobacteria were the most
abundant phyla in the fecal microbiota of healthy dogs [70, 86, 109]. However, Fusobacteria
(39.17%) were dominant, followed by Bacteroidetes (33.36%) and Firmicutes (15.81%) in healthy
adult Miniature Schnauzer dogs [86], while the abundances of Fusobacteria, Bacteroidetes and
Firmicutes were similar (approximately 30% each) in six Hound dogs [109].
Clostridia was the most predominant bacterial class in the dog fecal microbiota [109]. At the
genus level, Lactobacillus was the most predominant, followed by Bifidobacterium, Enterococcus,
Streptococcus and Pediococcus, in the dog fecal microbiota [110]. Many Lactobacillus spp. including
L. casei, L. salivarius, L. rhamnosus, L. mucosae, L. fermentum, L. reuteri, L. animalis, L. acidophilus
and L. johnsonii were the most frequently isolated ones from the feces. L. reuteri, L. animalis, and
L. johnsonii were the most predominant species in dogs [110–112]. Weissella confuse, Pediococcus
acidilactici, Enterococcus spp. and B. animalis ssp. lactis were also frequently isolated from dog feces
[111–113]. At the fungal kingdom-phylum level, Ascomycota, Basidiomycota, Glomeromycota,
and Zygomycota were detected in dog feces [109].
In the skin microbiota, the most predominant phyla and families were Proteobacteria and
Oxalobacteriaceae [87]. At the oral microbiota phylum level, Bacteroidetes (60%) was the most
predominant, followed by Proteobacteria (20.8%), Firmicutes (11.4%), Fusobacteria (4.7%) and
Spirochaetes (1.7%). At the genus level, the oral microbiota consisted of Porphyromonas (39.2%),
Fusobacterium (4.5%), Capnocytophaga (3.8%), Derxia (3.7%), Moraxella (3.3%) and Bergeyella
(2.7%) [88]. In the nasal microbiota of healthy dogs, Moraxella spp. was the most abundant species,
followed by Phyllobacterium spp., Staphylococcus spp., and Cardiobacteriaceae [90]. The most frequently
isolated bacteria from the dog’s vaginal tract were Lactobacillus, Escherichia coli and Staphylococcus
pseudointermedius [21, 91].
The feline GIT has different bacterial species than other animals. Helicobacter is known to reside
in the stomachs of cats [114]. For the microbiota composition of the GI (stomach, duodenum,
jejunum, ileum, and colon) contents, which were collected from 5 healthy felines, Firmicutes (68%)
predominated, followed by Proteobacteria (14%), Bacteroidetes (10%), Fusobacteria (5%) and
Actinobacteria (4%). At the order level, Clostridiales (54%) prevailed, followed by Lactobacillales,
Bacteroidales, Campylobacterales, and Fusobacteriales [37]. Based on several studies, it is known
that Bacteroides spp., Clostridium spp., Enterococcus spp., Streptococcus spp., Fusobacteria spp., and
Eubacteria spp. are present in the small intestines of felines [69,115]. Representative lactic acid
https://doi.org/10.5187/jast.2022.e8 https://www.ejast.org | 205Probiotics and the gut microbiome in companion animals
bacteria present in the GIT of felines include L. acidophilus, L. salivarius, L. johnsonii, L. reuteri
and L. sakei, which are typical intestinal lactic acid bacteria found in animals, including humans,
although the amount varies by individual [21,37]. The major phyla in the feline fecal microbiota
were Firmicutes, Bacteroidetes, Actinobacteria and Proteobacteria [69,109,116]. These four
phyla make up more than 99% of the fecal microbiota [69]. Handl et al. [109] reported that
Firmicutes was the most prevalent phylum in the fecal microbiota, followed by Bacteroidetes and
Actinobacteria; however, Tun et al. [116] reported that Bacteroidetes was the most predominant
phylum, followed by Firmicutes and Proteobacteria. Bacteroides, Fusobacterium, and Prevotella were
the most predominant genera in the feline fecal microbiota, which indicated that these genera
play a major role in the feline intestine [117]. Fungi, archaea and viruses compose a minor part
of intestinal microbial communities. Ascomycota was the only phylum of fungi detected in cats
[109,116].
Malassezia spp. were the most prevalent fungi in feline skin mycobiota. M. restricta and M.
globosa were the most predominant fungal species in all cat breeds [96]. M. pachydermatis is known
as a yeast that is present in the skin microbiome, yet it can also act as a pathogen that can cause
dermatitis [118]. The phylum level of the oral microbiota is generally conserved between cats. These
phyla are predominated by Proteobacteria (75.2%), followed by Bacteroidetes (9.3%), Firmicutes
(6.7%), SR1 (2.7%), Spirochaetes (1.8%), Fusobacteria (1.3%), and Actinobacteria (0.6%) [89]. The
composition of the canine and feline microbiota is shown in Figs. 1 and 2.
Fig. 1. The dynamic community of nasal, oral and gut microbiota in canines ([21], [37], [38], [70], [73], [86]–
[88], [90], [91], [107]–[113]).
206 | https://www.ejast.org https://doi.org/10.5187/jast.2022.e8Lee et al.
Fig. 2. The microbial community of feline nasal, oral and gut environments ([37],[69],[89], [114],[115],[117]).
SAFETY ISSUES OF PROBIOTICS AND THE GUT MICRO-
BIOME IN COMPANION ANIMALS
The Food and Agriculture Organization of the United Nations and World Health Organization
(FAO/WHO) defined probiotics as “live microorganisms, which when administered in adequate
amounts, confer a health benefit on the host” [9]. Tremendous scientific evidence for the efficacy of
probiotic candidates has been available for decades, but insufficient information about their safety
is available. While known to be safe in general, a few adverse effects associated with probiotics
use have been documented in patients [119,120]. Moreover, there is a lack of information on the
inherent characteristics of each probiotic strain that may be associated with health risks [121].
The European Food Safety Authority (EFSA) recommended the qualified presumption of
safety (QPS) status for microorganisms used in feed and food production in 2003. Based on the
QPS guidelines, microbes that produce toxins or possess virulence factors that may contribute to
their pathogenicity cannot be used as probiotics. In addition, it must be ensured that there are no
acquired genes encoding antimicrobial resistance (AMR). The existence of knowledge, including
a history of use, ecology, industrial application, clinical reports, and a public database, is considered
important evidence for evaluating the safety of microbial species [122]. QPS list includes several
taxonomic units for bacteria, yeasts, and viruses [123], of which Lactobacillus and Bifidobacterium
are representative because their reasonable certainty of no harm has been supported by an extensive
record of safe use [124].
With the recent focus on the beneficial effects of probiotics in companion animals and their
relationship to gut microflora and health, probiotic products are being increasingly marketed in the
form of feed additives, dietary supplements, and probiotic-containing foods [125]. Although there
have been no reports of adverse events when probiotics are administered to small animals, safety
concerns remain to be addressed [126]. The microorganisms used in feed additives require safety
verification for target animals, manufacturers, and owners/consumers. In particular, Enterococcus,
known as canine and feline intestinal commensal bacteria, have been used as probiotics for small
animals, but their use is restricted in some countries because of the risk of host infection by AMR
gene transfer [125,127]. Rinkinen [128] demonstrated that some Enterococcus faecium strains
https://doi.org/10.5187/jast.2022.e8 https://www.ejast.org | 207Probiotics and the gut microbiome in companion animals
promoted the adherence of the zoonotic pathogen Campylobacter jejuni in the intestines of canines.
Notably, the administration of E. faecium strain SF68 (deposited as strain NCIMB 10415), which
originated from infant feces in 1968, reduced the occurrence of diarrhea in dogs and cats housed
in animal shelters [45], and it has been verified that the strain may not cause any safety concerns
for companion animals and their owners [129–131]. Due to these conflicting outcomes, stringent
safety evaluations are required in a strain-specific manner with regard to probiotic use.
Host specificity is considered an important criterion for selecting probiotic candidates, primarily
due to differences in physiological structure, immune systems, and microbial composition
[132,133]. However, most commercial probiotics used as feed additives originate from humans
and are verified by human-based methods and criteria [134]. The clinical results from Weese
and Anderson [135] showed that Lactobacillus rhamnosus GG, a commercial probiotic strain
isolated from a healthy human GIT, may not be suitable for use in canines because of its short
persistence. In addition, in an in vitro test, probiotic strains of canine origin inhibited the adhesion
of enterotoxigenic Clostridium perfringens to canine jejunal chyme more efficiently than non-canine
strains [128]. Recent studies have focused on strains isolated from the intestines of healthy dogs
and cats to demonstrate their impact on pathogen inhibition, attenuation of inflammatory status,
and modulation of the gut microbiome [136,137]. Host specificity has been discussed with respect
to probiotic efficacy in most publications but must also be addressed from a safety perspective.
Furthermore, clinical outcomes for the safe use of probiotics in target animal species should be
documented. Given the host specificity and broad diversity of potential probiotic candidates for
small animals, the availability of novel species with no record of use requires attention [133]. For
a complete characterization, documentation of genetic and biochemical properties and long-term
clinical trials are needed. Additional efforts are required to standardize safety assessment methods
for novel species to be considered QPS or as having a generally recognized as safe status based on
the opinions of regulatory bodies and expert panels [138].
Quality control issues associated with probiotics relate to products intended for animals as
well as humans [21]. The global market for probiotics for companion animals is growing, but
insufficient quality control regulations create serious problems for the safety of consumers and
target animals. Some investigators have disclosed that many commercial animal probiotics or
pet foods that claim to contain probiotics did not contain microbial species listed on the label or
even contain other species. Moreover, bacterial viability, a key concept to stipulate probiotics, was
inconsistent with labeled values expressed in colony-forming units [139–141]. The current low level
of quality control may lead to exposure to unknown health risks not only for the animals but also
for the owners and the environment. Recent advances in meta-omics technologies (metagenomics,
metatranscriptomics, metaproteomics, and metabolomics) have promoted a correct evaluation of
the quality of probiotic products. For the multistrain product VSL#3, Mora and colleagues [142]
successfully identified microbial taxa with metagenomics and viability by flow cytometry and
confirmed reproducibility using metaproteomics. Metagenomic approaches have also enabled the
analysis of genes related to safety concerns in a culture-independent manner. Stringent oversight
by regulatory bodies, high manufacturer awareness, and the development of rigorous evaluation
methods by researchers are needed for the safe selection and production of probiotic candidates
intended for both companion animal and human use.
CONCLUSION
Although not enough research has been conducted on the probiotics and gut microbiome thus
far, because the number of companion animal people and the companion animal market are
208 | https://www.ejast.org https://doi.org/10.5187/jast.2022.e8Lee et al.
growing, research related to the companion animal microbiome is also growing. Recently, in-depth
research has been performed to identify the functionality of probiotics and the gut microbiome of
companion animals and to make them with various materials, ranging from feed, snacks, supplies,
and treatments for the diseases of companion animals. The current evidence suggests that specific
probiotic strains and/or their defined combinations may be useful in canine and feline nutrition,
therapy and care. However, probiotics and the gut microbiota used in the present study are of
human origin; thus, the companion animal-specific health benefits are not unclear. Therefore, the
most important step is to secure pet-originated microorganisms for their health claims. Moreover,
detailed in vivo designs and trials using companion animals are needed to identify and characterize
newly isolated pet-originated microbiomes with an impact on health maintenance in both dogs and
cats. Corroborations of these health-promoting effects and microbiological safety issues should be
assessed regarding potential probiotics and the gut microbiome for animal health and welfare.
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