An October 2001 report by the  World Health Organization  (WHO) defines probiotics as "live microorganisms which when administered in adequate amounts confer a health benefit on the host." [ 12 ] [ 13 ]  Following this definition, a  working group  convened by the  Food and Agriculture Organization  (FAO)/WHO in May 2002 issued the  Guidelines for the Evaluation of Probiotics in Food . [ 14 ]  A consensus definition of the term  probiotics , based on available information and scientific evidence, was adopted after the aforementioned joint expert consultation between the  FAO  of the  United Nations  and the WHO. This effort was accompanied by local governmental and supra-governmental regulatory bodies' requirements to better characterize health claims substantiations. [ citation needed ]


That first global effort was further developed in 2010; two expert groups of academic scientists and industry representatives made recommendations for the evaluation and validation of probiotic health claims. [ 15 ] [ 16 ]  The same principles emerged from those two groups as were expressed in the "Guidelines" of FAO/WHO in 2002. This definition, though widely adopted, is not acceptable to the  European Food Safety  Authority because it embeds a health claim that is not measurable. [ 7 ]


A group of scientific experts assembled in Canada in October 2013 to discuss the scope and appropriate use of the term "probiotic", adjusting the definition to be "live microorganisms that, when administered in adequate amounts, confer a health benefit on the host." [ 17 ]


Live probiotic  cultures  are part of  fermented dairy products , other  fermented  foods, and probiotic-fortified foods. [ 18 ]


Lactic acid bacteria  (LAB), which are food fermenting bacteria, have the ability to prevent food spoilage and can improve the nutritive value of the foods they inhabit. Acid fermentation (as well as salting), remains one of the most practical methods of preservation of fresh vegetables, cereal gruels, and milk-cereal mixtures due to its low cost and energy requirements. [ 19 ]


Fermented products  that contain lactic acid bacteria include vegetables such as  pickled vegetables , [ 20 ]   kimchi , [ 20 ] [ 21 ]   pao cai , [ 22 ]  and  sauerkraut ; [ 23 ]   sourdough bread  or bread-like products made without  wheat  or  rye flour , amino acid/peptide meat-flavored sauces and pastes produced by fermentation of cereals and legumes; fermented cereal-fish-shrimp mixtures and fermented meats; [ 19 ]  soy products such as  tempeh , [ 24 ]   miso , [ 25 ]  and  soy sauce ; [ 26 ]  dairy products such as  yogurt ,  kefir , [ 27 ]   buttermilk ; [ 28 ]  and non-dairy products such as  bee pollen . [ 29 ]


More precisely, sauerkraut contains the bacteria  Leuconostoc mesenteroides ,  Lactobacillus plantarum , Pediococcus pentosaceus,  Lactobacillus brevis , Leuconostoc citreum, Leuconostoc argentinum,  Lactobacillus paraplantarum , Lactobacillus coryniformis , and  Weissella  spp. [ 30 ]  Kimchi contains the bacteria  Leuconostoc  spp.,  Weissella  spp., and  Lactobacillus  spp. Pao cai contains  L. pentosus, L. plantarum , Leuconostoc mesenteroides , L. brevis, L. lactis , and  L. fermentum .
A list of many other bacteria found in several Asian fermented fruits and vegetables also is available. [ 31 ] [ 32 ]  Kefir contains  Lactobacillus acidophilus ,  Bifidobacterium bifidum ,  Streptococcus thermophilus ,  Lactobacillus delbrueckii  subsp.  bulgaricus, Lactobacillus helveticus,  Lactobacillus kefiranofaciens , Lactococcus lactis , and  Leuconostoc  species. [ 33 ] [ 34 ]  Buttermilk contains either  Lactococcus lactis  or  L. bulgaricus . Other acidic bacteria, said to be probiotic, [ 35 ] [ 36 ]  can be found in  kombucha , including  Gluconacetobacter xylinus , [ 37 ] [ 38 ]   Zygosaccharomyces  sp.,  Acetobacter pasteurianus,  Acetobacter aceti , and  Gluconobacter oxydans . [ 39 ]


It is incorrect to think that a higher colony forming units (CFU) count corresponds to greater efficacy - rather, probiotic efficacy is strain- and disease specific. [ 40 ]


The manipulation of the gut microbiota is complex and may cause bacteria-host interactions. [ 5 ]  Though probiotics are considered safe, some have concerns about their safety in certain cases. [ 5 ] [ 41 ]  Some people, such as those with  immunodeficiency ,  short bowel syndrome ,  central venous catheters , and  cardiac valve disease , and premature infants, may be at higher risk for adverse events. [ 3 ]  In severely ill people with  inflammatory bowel disease , a risk exists for the passage of viable bacteria from the gastrointestinal tract to the internal organs (bacterial translocation) as a consequence of  bacteremia , which can cause adverse health consequences. [ 5 ]  Rarely, consumption of probiotics by children with lowered immune system function or who are already critically ill may result in bacteremia or  fungemia  (i.e., bacteria or fungi in the blood), which can lead to  sepsis , a potentially fatal disease. [ 4 ]


Probiotic supplements typically contain between one and ten billion  colony-forming units  (CFUs) per dose. [ 42 ]  A higher number of CFUs does not provide additional probiotic effects, but may have unintended consequences of causing digestive discomfort, such as bloating, gas, and diarrhea. [ 42 ] [ 43 ]


Lactobacillus  species have been suggested to contribute to obesity in humans, but no evidence of this relationship has been found. [ 44 ]


In 2015, the global retail market value for probiotics was US$41 billion, including sales of probiotic  supplements , fermented milk products, and yogurt, which alone accounted for 75% of total consumption. [ 45 ]  Innovation in probiotic products in 2015 was mainly from supplements, which produced US$4 billion and was projected to grow 37% globally by 2020. [ 45 ]  Consumption of yogurt products in China has increased by 20% per year since 2014. [ 46 ]


As of 2019 [update] , the  European Food Safety Authority  has rejected all petitions by commercial manufacturers for health claims on probiotic products in Europe due to insufficient evidence for a  cause-and-effect mechanism  for benefit,  thus inconclusive proof of effectiveness. [ 7 ] [ 8 ] [ 11 ]  The  European Commission  placed a ban on putting the word "probiotic" on the packaging of products because such labeling misleads consumers to believe a health benefit is provided by the product when no scientific proof exists to demonstrate that health effect. [ 7 ] [ 47 ] [ 48 ] [ 49 ]


In the United States, the  Food and Drug Administration  (FDA) and  Federal Trade Commission  (FTC) have issued warning letters and imposed punishment on various manufacturers of probiotic products whose labels claim to treat a disease or condition. [ 10 ] [ 50 ] [ 51 ]   Food product labeling  requires language approved by the FDA, so probiotic manufacturers have received warning letters for making disease or treatment claims. [ 10 ] [ 51 ]  The FTC has taken punitive actions, including a US$21 million fine coordinated by 39 different state governments against a major probiotic manufacturer for deceptive advertising and exaggerated claims of health benefits for yogurt and probiotic dairy drink. [ 50 ]


In  Vietnam , the Vietnam Food Administration (VFA) under the  Ministry of Health , in collaboration with other relevant authorities, oversees and addresses violations related to probiotic products. This includes issuing warnings, imposing administrative penalties, [ 52 ]  demanding product recalls [ 53 ] [ 54 ]  and coordinating with other agencies. [ 55 ] [ 56 ]


The  National Yogurt Association  (NYA) of the United States gives a "Live & Active Cultures Seal" to refrigerated yogurt products that contain 100 million cells per gram, or frozen yogurt products that contain 10 million cells per gram at the time of manufacture. [ 57 ]  In 2002, the FDA and WHO recommended that "the minimum viable numbers of each probiotic strain at the end of the shelf-life" be reported on labeling, [ 58 ]  but most companies that give a number report the viable cell count at the date of manufacture, a number that could be much higher than that which exists at consumption. [ 59 ]  Because of the variability in storage conditions and time before eating, exactly how many active culture cells remain at the time of consumption is difficult to determine. The survival of probiotics was strongly dependent on the storage temperature and remarkable viability loss occurred in room temperature compared to refrigerated storage. [ 60 ]


Probiotics have received renewed attention in the 21st century from product manufacturers, research studies, and consumers. Their history can be traced to the first use of cheese and fermented products, which were well-known to the  Greeks  and  Romans  who recommended their consumption. [ 61 ]  The  fermentation  of dairy foods represents one of the oldest techniques for  food preservation . [ 62 ]


The original modern hypothesis of the positive role played by certain bacteria was first introduced by  Russian  scientist and  Nobel laureate   Élie Metchnikoff , who in 1907 suggested that it would be possible to modify the  gut microbiota  and to replace harmful microbes with useful microbes. [ 63 ]  Metchnikoff, at that time a professor at the  Pasteur Institute  in  Paris , proposed the hypothesis that the  aging  process results from the activity of  putrefactive  ( proteolytic ) microbes producing toxic substances in the  large bowel . Proteolytic bacteria such as  clostridia , which are part of the normal gut microbiota, produce toxic substances including  phenols ,  indols , and  ammonia  from the  digestion  of  proteins . According to  Metchnikoff , these compounds were responsible for what he called "intestinal  autointoxication ", which would cause the physical changes associated with old age. [ 64 ]


At that time,  milk fermented  with  lactobacillales  were known to inhibit the growth of proteolytic bacteria because of the low  pH  produced by the fermentation of  lactose . Metchnikoff had also observed that certain rural populations in Europe, for example in Bulgaria and the Russian steppes, who lived largely on milk fermented by lactic-acid bacteria, were exceptionally long-lived. Based on these observations, Metchnikoff proposed that consumption of fermented milk would "seed" the  intestine  with harmless lactic-acid bacteria and decrease the intestinal pH, and that this would suppress the growth of proteolytic bacteria. Metchnikoff himself introduced in his diet  sour milk  fermented with the bacteria he called "Bulgarian Bacillus" and believed his health benefited. Friends in Paris soon followed his example and physicians began prescribing the sour-milk diet for their patients. [ 65 ]


Bifidobacteria  was first isolated from a breastfed infant by Henry Tissier, who also worked at the  Pasteur Institute . The isolated bacterium named  Bacillus bifidus communis [ 66 ]  was later renamed to the genus  Bifidobacterium . [ 67 ]  Tissier found that bifidobacteria are dominant in the gut microbiota of  breast-fed  babies and he observed clinical benefits from treating infant diarrhea with bifidobacteria.


During an outbreak of  shigellosis  in 1917, German professor Alfred Nissle isolated a strain of  Escherichia coli  from the feces of a soldier who was not affected by the disease. [ 68 ]  Methods of treating infectious diseases were needed at that time when antibiotics were not yet available, and Nissle used the  E. coli  Nissle 1917 strain  in acute gastrointestinal infectious  salmonellosis  and  shigellosis . [ 69 ]


In 1920, Rettger and Cheplin reported that Metchnikoff's "Bulgarian Bacillus", later called  Lactobacillus delbrueckii  subsp.  bulgaricus , could not live in the human intestine. [ 70 ] [ non-primary source needed ]  They conducted experiments involving rats and humans volunteers, feeding them with  Lactobacillus acidophilus . They observed the disappearance of the pathogenic protist  Balantidium coli  as well as of other gas-producing bacteria. [ 70 ]  Rettger further explored the possibilities of  L. acidophilus , and reasoned that bacteria originating from the gut were more likely to produce the desired effect in this environment. In 1935, certain strains of  L. acidophilus  were found very active when implanted in the human digestive tract. [ 71 ] [ non-primary source needed ]


Contrasting antibiotics, probiotics were defined as microbially derived factors that stimulate the growth of other microorganisms. In 1989, Roy Fuller suggested a definition of probiotics that have been widely used: "A live microbial feed supplement which beneficially affects the host animal by improving its intestinal microbial balance." [ 72 ]  Fuller's definition emphasizes the requirement of viability for probiotics and introduces the aspect of a beneficial effect on the host.


The term "probiotic" originally referred to microorganisms that have effects on other microorganisms. [ 73 ]  The concept of probiotics involved the notion that substances secreted by one microorganism stimulated the growth of another microorganism. The term was used again [ 74 ]  to describe tissue extracts that stimulated microbial growth. The term probiotics was taken up by Parker, [ 75 ]  who defined the concept as, "Organisms and substances that have a beneficial effect on the host animal by contributing to its intestinal microbial balance." Later, the definition was greatly improved by Fuller, [ 72 ]  whose explanation was very close to the definition used today. Fuller described probiotics as a "live microbial feed supplement which beneficially affects the host animal by improving its intestinal microbial balance." He stressed two important claims for probiotics: the viable nature of probiotics and the capacity to help with intestinal balance.


In the following decades, intestinal lactic-acid bacterial species with alleged health-beneficial properties were introduced as probiotics, including  Lactobacillus rhamnosus ,  Lactobacillus casei , and  Lactobacillus johnsonii . [ 76 ]


Some literature gives the word a full Greek  etymology , [ 77 ] [ 78 ]  but it appears to be a composite of the Latin preposition  pro , meaning 'for', and the Greek adjective βιωτικός ( biōtikos ), meaning 'fit for life, lively', [ 79 ]  the latter deriving from the noun βίος ( bios ), meaning 'life'. [ 80 ]  The term contrasts etymologically with the term  antibiotic , although it is not a complete  antonym . The related term  prebiotic  comes from the Latin  prae , meaning 'before', and refers to a substance that is not digested, but rather may be  fermented  to promote the growth of beneficial intestinal microorganisms. [ 81 ]


As food products or dietary supplements, probiotics are under preliminary research to evaluate if they provide any effect on health. [ 2 ] [ 7 ] [ 82 ]  In all cases proposed as health claims to the  European Food Safety Authority , the scientific evidence remains insufficient to prove a cause-and-effect relationship between consumption of probiotic products and any health benefit. [ 7 ] [ 83 ]  There is no scientific basis for extrapolating an effect from a tested strain to an untested strain. [ 2 ] [ 84 ] [ 85 ]  Improved health through gut flora modulation appears to be directly related to long-term dietary changes. [ 7 ] [ 86 ]  Claims that some lactobacilli may contribute to  weight gain  in some humans [ 87 ] [ 88 ]  remain controversial. [ 89 ]


There is inconsistency in the results of different groups of 3488 children as reported in a Cochrane review. [ 90 ]  Also, it shows no significant difference regarding the adverse effects between probiotic and the other comparators. [ 90 ]


Only limited, low-quality evidence exists to indicate that probiotics are helpful for treating people with  milk allergy . [ 91 ]  A 2015 review showed low-quality evidence that probiotics given directly to infants with  eczema , or in infants whose mothers used probiotics during the  last trimester of pregnancy  and breastfeeding, had lower risk of eczema. [ 92 ]


It is unclear whether probiotic supplementation helps with childhood  asthma , as the quality of research evidence is low. [ 93 ]


Antibiotics are a common treatment for children, with 11% to 40% of antibiotic-treated children developing  diarrhea . [ 94 ]   Antibiotic-associated diarrhea  (AAD) results from an imbalance in the colonic microbiota caused by antibiotic therapy. [ 94 ]  These microbial community alterations result in changes in  carbohydrate metabolism , with decreased  short-chain fatty acid  absorption and osmotic diarrhea as a result. A 2015  Cochrane review  concluded that a protective effect of some probiotics existed for AAD in children. [ 94 ]  The known risks of using probiotics for treating  Clostridioides difficile  outweighs the uncertain benefits. [ 95 ]


Probiotic treatment might reduce the incidence and severity of AAD as indicated in several  meta-analyses . [ 96 ] [ 97 ] [ 98 ]  For example, treatment with probiotic formulations including   L. rhamnosus  may reduce the risk of AAD, improve stool consistency during antibiotic therapy, and enhance the immune response after vaccination. [ 99 ]


The potential efficacy of probiotics to treat AAD depends on the probiotic strains and dosage. [ 100 ] [ 101 ]  One review recommended for children  L. rhamnosus  or  Saccharomyces boulardii  at 5 to 40 billion colony-forming units/day, given the modest number needed to treat and the likelihood that adverse events are very rare. [ 94 ]  The same review stated that probiotic use should be avoided in pediatric populations at risk for  adverse events , such as severely debilitated or  immune-compromised  children. [ citation needed ]


Probiotic treatment of bacterial vaginosis is the application or ingestion of  bacterial species found in the healthy vagina  to cure the infection of bacteria causing  bacterial vaginosis . This treatment is based on the observation that 70% of healthy females have a group of bacteria in the genus  Lactobacillus  that dominate the population of organisms in the vagina. Specific strains of lactobacilli inhibit the growth of bacteria causing BV by producing H 2 O 2 , lactic acid, and/or bacteriocins, and/or inhibit the adherence of  Gardnerella vaginalis  to the vaginal epithelium, which prevents the infection from occurring in the vagina. [ 102 ]  Currently, the success of probiotic treatment has been mixed, since the use of probiotics to restore healthy populations of  Lactobacillus  has not been standardized.  Often, standard antibiotic treatment is used at the same time that probiotics are being tested. In addition, some groups of women respond to treatment based upon ethnicity, age, number of sexual partners, pregnancy, and the pathogens causing bacterial vaginosis. [ 103 ]  In 2013, researchers found that administration of  hydrogen peroxide -producing strains, such as  L. acidophilus  and  L. rhamnosus , were able to normalize vaginal pH and rebalance the  vaginal microbiota , preventing and alleviating bacterial vaginosis. [ 104 ]


As of 2017 [update] , only limited evidence indicated any direct link between  high blood pressure  and gut microbiota. [ 105 ]


A 2002 meta-analysis that included five double-blind trials examining the short-term (2–8 weeks) effects of a yogurt with probiotic strains on serum cholesterol levels found little effect of 8.5 mg/dL (0.22 mmol/L) (4% decrease) in  total cholesterol  concentration, and a decrease of 7.7 mg/dL (0.2 mmol/L) (5% decrease) in serum  LDL  concentration. [ 106 ]


A 2019 meta-analysis found low-quality evidence for probiotics having a small improvement in  depression  and  anxiety . [ 107 ]  A 2020 review found probiotics might improve depression, but more studies are needed. [ 108 ]


Some probiotics are suggested as a possible treatment for various forms of  gastroenteritis . [ 109 ]  As a treatment for infectious diarrhea, probiotics are of no benefit to people who have the condition for more than two days, and there is no evidence they lessen the duration of diarrhea overall. [ 110 ]


Probiotics are commonly given to breastfeeding mothers and their young children to prevent eczema ( dermatitis ), but no good evidence shows efficacy for this purpose. [ 111 ]  There is little evidence to support the use of probiotics to treat  atopic dermatitis , and some risk of  adverse effects . [ 112 ]  The  American Academy of Dermatology  stated: "The use of probiotics/prebiotics for the treatment of patients with established atopic dermatitis is not recommended due to inconsistent evidence". [ 113 ]


According to an umbrella review of meta-analyses of randomized controlled trials, probiotics supplementation reduces glucose homeostasis. This can be an effective therapy for lowering high blood sugar levels unless the body becomes  hypoglycemic ; caution and glucose monitoring are necessary to avoid this.  [ 114 ]


Some strains of lactic acid bacteria (LAB) may affect  Helicobacter pylori  infections (which may cause  peptic ulcers ) in adults when used in combination with standard medical treatments, but no standard in medical practice or regulatory approval exists for such treatment. [ 115 ]  The only peer-reviewed treatments for  H. pylori  to date all include various Antibiotic Regimens. [ 116 ]


Some strains of LAB may affect  pathogens  by means of  competitive inhibition  (i.e., by competing for growth) and some evidence suggests they may improve immune function by increasing the number of  IgA -producing plasma cells and increasing or improving  phagocytosis , as well as increasing the proportion of  T lymphocytes  and natural killer cells. [ 117 ] [ 118 ]  LAB products might aid in the treatment of acute diarrhea and possibly affect  rotavirus  infections in children and travelers' diarrhea in adults, [ 117 ] [ 118 ]  but no products are approved for such indications. There are weak evidence probiotics might lower the incidence of acute upper respiratory tract infections in adults, they were better than placebo or no treatment. [ 119 ]


Probiotics do not appear to change the risk of infection in older people. [ 120 ]


The use of oral probiotic supplements to modify the composition and behavior of the microbiome has been considered as a possible therapy for both induction and maintenance of remission in people with Crohn's disease and ulcerative colitis. A Cochrane review in 2020 did not find clear evidence of improved remission likelihood, nor lower adverse events, in people with Crohn's disease, following probiotic treatment.  [ 121 ]


For ulcerative colitis, there is low-certainty evidence that probiotic supplements may increase the probability of clinical remission.  [ 122 ]  People receiving probiotics were 73% more likely to experience disease remission and over 2x as likely to report improvement in symptoms compared to those receiving a placebo, with no clear difference in minor or serious adverse effects.  [ 122 ] Although there was no clear evidence of greater remission when probiotic supplements were compared with  5‐aminosalicylic acid  treatment as a  monotherapy , the likelihood of remission was 22% higher if probiotics were used in combination with 5-aminosalicylic acid therapy.  [ 122 ]  Whereas in people who are already in remission, it is unclear whether probiotics help to prevent future relapse, either as a monotherapy or  combination therapy .  [ 123 ]


Probiotics are under study for their potential to affect  irritable bowel syndrome , although uncertainty remains around which type of probiotic works best, and around the size of possible effect. [ 124 ] [ 125 ]


Several clinical studies provide evidence for the potential of probiotics to lower the risk of  necrotizing enterocolitis  and mortality in premature infants. One meta-analysis indicated that probiotics reduce these risks by more than 50% compared with controls but that further, large, high-quality trials were needed to inform policy and practice. [ 126 ]


A Cochrane systematic review found no good evidence that probiotics were of benefit in reducing the risk of  gestational diabetes , but good evidence that they increased the risk of  pre-eclampsia . For this reason, the use of probiotics in pregnancy was advised against. [ 127 ]


A 2017 review based on moderate to low-quality evidence suggests that probiotics may be helpful in relieving pain in the short term in children with recurrent abdominal pain, but the proper strain and dosage are not known. [ 128 ]


A clinical study investigating the impact of probiotics in relieving the signs and symptoms of  dry eye  revealed promising results for the ophthalmic formulation of  Latilactobacillus sakei , while the oral probiotic demonstrated no discernible benefits. [ 129 ]


There is limited evidence indicating probiotics are of benefit in the management of infection or inflammation of the  urinary tract . [ 130 ]  One literature review found  Lactobacillus  probiotic supplements appeared to increase vaginal lactobacilli levels, thus reducing the incidence of vaginal infections in otherwise healthy adult women. [ 131 ]


Supplements such as tablets, capsules, powders, and sachets containing bacteria have been studied.  However, probiotics taken orally can be destroyed by the acidic conditions of the stomach.  As of 2010, a number of  microencapsulation  techniques were being developed to address this problem. [ 132 ]


Preliminary research is evaluating the potential  physiological  effects of multiple probiotic strains, as opposed to a single strain. [ 133 ] [ 134 ]  As the human gut may contain tens of thousands of microbial species, one theory indicates that this diverse environment may benefit from consuming multiple probiotic strains, an effect that remains scientifically unconfirmed. [ citation needed ]


Only preliminary evidence exists for most probiotic health claims. Even for the most studied probiotic  strains , few have been sufficiently developed in basic and clinical research to warrant approval for  health claim  status by a regulatory agency such as the FDA or EFSA, and as of 2010 [update] , no claims had been approved by those two agencies. [ 7 ]  Some experts are skeptical about the efficacy of different probiotic strains and believe that not all subjects benefit from probiotics. [ 7 ] [ 135 ]


Multiple studies have shown that there is a significant difference in the survival rate of Lactobacillus and Bifidobacterium under refrigerated (4°C) and room temperature (25°C) storage conditions. At room temperature (25±1°C), the number of  probiotics decreased by 5 to 6 logarithmic units (down to 1/100,000) after 90 days of storage. In contrast, no significant change in the number of probiotics was observed under refrigerated conditions (4 ± 1°C). [ 136 ]


First, probiotics must be alive when administered. [ 72 ] [ 137 ] [ 138 ]  One of the concerns throughout the  scientific literature  resides in the viability and reproducibility on a large scale of observed results for specific studies, as well as the viability and stability during use and storage, and finally the ability to survive in stomach acids and then in the intestinal ecosystem. [ 7 ] [ failed verification ]


Second, probiotics must have undergone controlled evaluation to document  health benefits  in the target host. Only products that contain live organisms shown in reproducible human studies to confer a health benefit may claim to be probiotic. [ 7 ] [ 139 ] [ 140 ]  The correct definition of health benefit, backed with solid scientific evidence, is a strong element for the proper identification and assessment of the effect of a probiotic. This aspect is a challenge for scientific and industrial investigations because several difficulties arise, such as variability in the site for probiotic use (oral, vaginal, intestinal) and mode of application. [ 72 ]


Third, the probiotic candidate must be a taxonomically defined microbe or combination of microbes ( genus ,  species , and strain level). It is commonly admitted that most effects of probiotics are strain-specific and cannot be extended to other probiotics of the same genus or species. [ 137 ]  This calls for precise identification of the strain, i.e.  genotypic  and  phenotypic  characterization of the tested microorganism. [ 15 ]


Fourth, probiotics must be safe for their intended use. The 2002 FAO/WHO guidelines recommend that, though bacteria may be generally recognized as safe (GRAS), the safety of the potential probiotic be assessed by the  minimum  required tests:  [ 141 ]


In Europe, EFSA adopted a premarket system for the safety assessment of microbial species used in food and feed productions to set priorities for the need for risk assessment. The assessment is made for certain microorganisms; if the result is favorable, it leads to "Qualified Presumption of Safety"  status. [ 143 ]


Psychobiotics  is a term used in preliminary research to refer to  live bacteria  that, when ingested in appropriate amounts, might confer a  mental health  benefit by affecting  microbiota  of the host organism. [ 1 ]  Whether bacteria might play a role in the  gut-brain axis  is under research. A 2020 literature review suggests that the consumption of psychobiotics could be considered as a viable option to restore mental health [ 2 ]  although lacking  randomized controlled trials  on clear mental health outcomes in humans. [ 3 ] [ 4 ]


In experimental probiotic psychobiotics, the bacteria most commonly used are  gram-positive bacteria , such as  Bifidobacterium  and  Lactobacillus  families, as these do not contain  lipopolysaccharide  chains, reducing the likelihood of an  immunological response . [ 1 ]   Prebiotics  are substances, such as  fructans  and  oligosaccharides , that induce the growth or activity of beneficial  microorganisms , such as bacteria on being  fermented  in the gut. [ 1 ] [ 5 ]  Multiple bacterial species contained in a single probiotic broth is known as a polybiotic. [ 6 ]


Psychobiotics are probiotics that influence the gut-brain axis, the bidirectional communication network linking the gastrointestinal system and the brain. By positively modulating the gut microbiota, psychobiotics can impact the production of neurotransmitters, reduce inflammation, and enhance neural signaling, which may improve mental health and alleviate symptoms of neurological disorders. This highlights their potential as a therapeutic tool for maintaining a healthy gut-brain connection.


A 2021 review showed that treating anxiety in young people with psychobiotics had no significant effect. [ 7 ]  This study highlights the gut microbiome's role in brain function and mental health is a growing research area, particularly during adolescence, a critical period for gut-brain axis development. This study systematically reviewed and analyzed the effects of psychobiotic interventions on anxiety in youth (ages 10–24). Among 5416 studies identified, 14 were reviewed, and 10 included in the meta-analysis, involving 617 participants. The results showed minimal efficacy of psychobiotics in reducing anxiety, with a pooled standard mean difference of −0.03, indicating no significant effect. Despite limited evidence, future research focusing on mechanisms, causality, and youth perspectives is essential for advancing psychobiotic use in mental health management.


Another  2022  article highlights how the nervous system disorders affect millions globally each year, sparking interest in the potential of probiotics to support the gut-brain axis alongside traditional treatments. Psychobiotics, a type of probiotic, have shown promise in improving gut microbiota and alleviating symptoms of central nervous system (CNS) disorders. This study reviewed their impact on conditions like autism spectrum disorders, Parkinson's disease, multiple sclerosis, depression, and insomnia, among others. The findings suggest that psychobiotic-containing functional foods could be an effective tool for improving mental health and managing CNS disorders.


There is a need for more diverse human studies, mainly because those that exist have contradictory outcomes. [ 3 ] [ 4 ] [ 7 ]


Several species of bacteria have been used in probiotic psychobiotic research: [ 6 ] [ 8 ]


Putrefaction  is the fifth  stage of death , following  pallor mortis ,  livor mortis ,  algor mortis , and  rigor mortis . This process references the breaking down of a body of an  animal   post-mortem . In broad terms, it can be viewed as the decomposition of  proteins , and the eventual breakdown of the cohesiveness between tissues, and the liquefaction of most organs. This is caused by the decomposition of organic matter by bacterial or fungal digestion, which causes the release of gases that infiltrate the body's tissues, and leads to the deterioration of the tissues and organs.
The approximate time it takes putrefaction to occur is dependent on various factors. Internal factors that affect the rate of putrefaction include the age at which death has occurred, the overall structure and condition of the body, the cause of death, and external injuries arising before or after death. External factors include environmental temperature, moisture and air exposure, clothing, burial factors, and light exposure.  Body farms  are facilities that study the way various factors affect the putrefaction process.


The first signs of putrefaction are signified by a greenish discoloration on the outside of the skin, on the abdominal wall corresponding to where the large intestine begins, as well as under the surface of the liver.


Certain substances, such as  carbolic acid ,  arsenic ,  strychnine , and  zinc chloride , can be used to delay the process of putrefaction in various ways based on their chemical make up.


In  thermodynamic  terms, all organic tissues are composed of chemical energy, which, when not maintained by the  constant biochemical maintenance  of the living organism, begin to chemically break down due to the reaction with water into  amino acids , known as  hydrolysis . The breakdown of the proteins of a decomposing body is a  spontaneous process .  Protein hydrolysis  is accelerated as the  anaerobic bacteria  of the  digestive tract  consume, digest, and excrete the cellular proteins of the body.


The bacterial digestion of the cellular proteins weakens the tissues of the body. As the proteins are continuously broken down to smaller components, the bacteria excrete gases and  organic compounds , such as the  functional-group   amines   putrescine  (from  ornithine ) and  cadaverine  (from  lysine ), which carry the noxious odor of rotten flesh. Initially, the gases of putrefaction are constrained within the body cavities, but eventually diffuse through the adjacent tissues, and then into the  circulatory system . Once in the blood vessels, the putrid gases infiltrate and diffuse to other parts of the body and the limbs.


The visual result of gaseous tissue-infiltration is notable bloating of the torso and limbs. The increased internal pressure of the continually rising volume of gas further stresses, weakens, and separates the tissues constraining the gas. In the course of putrefaction, the skin tissues of the body eventually rupture and release the bacterial gas. As the anaerobic bacteria continue consuming, digesting, and excreting the tissue proteins, the body's decomposition progresses to the stage of  skeletonization . This continued consumption also results in the production of  ethanol  by the bacteria, which can make it difficult to determine the  blood alcohol content  (BAC) in autopsies, particularly in bodies recovered from water. [ 1 ]


Generally, the term  decomposition  encompasses the biochemical processes that occur from the physical death of the person (or animal) until the skeletonization of the body. Putrefaction is one of seven stages of  decomposition ; as such, the term  putrescible  identifies all organic matter (animal and human) that is  biochemically  subject to putrefaction. In the matter of death by poisoning, the putrefaction of the body is chemically delayed by poisons such as  antimony ,  arsenic ,  carbolic acid  (phenol),  nux vomica  (plant),  strychnine  (pesticide), and  zinc chloride .


The rough timeline of events during the putrefaction stage is as follows:


Order of organs' decomposition in the body: [ 2 ]


The rate of putrefaction is greatest in air, followed by water, soil, and earth. The exact rate of putrefaction is dependent upon many factors such as weather, exposure and location. Thus, refrigeration at a morgue or funeral home can retard the process, allowing for burial in three days or so following death without  embalming .  The rate increases dramatically in tropical climates. The first external sign of putrefaction in a body lying in air is usually a greenish discoloration of the skin over the region of the  cecum , which appears in 12–24 hours. The first internal sign is usually a greenish discoloration on the undersurface of the liver.


Various factors affect the rate of putrefaction. [ 3 ] [ 4 ] [ 5 ]


Environmental temperature: Decomposition is accelerated by high atmospheric or environmental temperature, with putrefaction speed optimized between 21 °C (70 °F) and 38 °C (100 °F), further sped along by high levels of humidity. This optimal temperature assists in the chemical breakdown of the tissue and promotes microorganism growth. Decomposition nearly stops below 0 °C (32 °F) or above 48 °C (118 °F).


Moisture and air exposure: Putrefaction is ordinarily slowed by the body being submerged in water, due to diminished exposure to air. Air exposure and moisture can both contribute to the introduction and growth of microorganisms, speeding degradation. In a hot and dry environment, the body can undergo a process called mummification where the body is completely dehydrated and bacterial decay is inhibited.


Clothing: Loose-fitting clothing can speed up the rate of putrefaction, as it helps to retain body heat. Tight-fitting clothing can delay the process by cutting off blood supply to tissues and eliminating nutrients for bacteria to feed on.


Manner of burial: Speedy burial can slow putrefaction. Bodies within deep graves tend to decompose more slowly due to the diminished influences of changes in temperature. The composition of graves can also be a significant contributing factor, with dense, clay-like soil tending to speed putrefaction while dry and sandy soil slows it.


Light exposure: Light can also contribute indirectly, as flies and insects prefer to lay eggs in areas of the body not exposed to light, such as the crevices formed by the eyelids and nostrils. [ 3 ]


Age at time of death: Stillborn fetuses and infants putrefy slowly due to their sterility. Otherwise, however, younger people generally putrefy more quickly than older people. [ citation needed ]


Condition of the body: A body with a greater fat percentage and less lean body mass will have a faster rate of putrefaction, as fat retains more heat and it carries a larger amount of fluid in the tissues. [ 5 ]


Cause of death: The cause of death has a direct relationship to putrefaction speed, with bodies that died from acute violence or accident generally putrefying slower than those that died from infectious diseases. Certain poisons, such as  potassium cyanide  or  strychnine , may also delay putrefaction, while chronic alcoholism and cocaine use will speed it. [ citation needed ]


External injuries: Antemortem or postmortem injuries can speed putrefaction as injured areas can be more susceptible to invasion by bacteria. [ citation needed ]


Certain poisonous substances to the body can delay the process of putrefaction. They include:


Embalming  is the process of preserving human remains by delaying decomposition. This is acquired through the use of embalming fluid, which is a mixture of formaldehyde, methanol, and various other solvents. The most common reasons to preserve the body are for viewing purposes at a funeral, for above-ground interment or distant transportation of the deceased, and for medical or religious practices.


Body farms  subject donated cadavers to various environmental conditions to study the process of human decomposition. [ 7 ]  These include The University of Tennessee's Forensic Anthropologic Facility, Western Carolina Universities Osteology Research Station (FOREST), Texas State University's Forensic Anthropology Research Facility (FARF), Sam Houston State University's Southeast Texas Applied Forensic Science Facility (STAFS), Southern Illinois University's Complex for Forensic Anthropology Research, and Colorado Mesa University's Forensic Investigation Research Station. The Australian Facility for Taphonomic Experimental Research, near  Sydney , is the first body farm located outside of the United States  [ 8 ]  In the United Kingdom there are several facilities which, instead of using human remains or cadavers, use dead pigs to study the decomposition process. Pigs are less likely to have infectious diseases than human cadavers, and are more readily available without any concern for ethical issues, but a human body farm is still highly sought after for further research. [ 9 ]  Each body farm is unique in its environmental make-up, giving researchers a broader knowledge, and allowing research into how different environmental factors can affect the rate of decomposition significantly such as humidity, sun exposure, rain or snow, altitude level and more.


In  alchemy , putrefaction is the same as  fermentation , whereby a substance is allowed to rot or decompose undisturbed.  In some cases, the commencement of the process is facilitated with a small sample of the desired material to act as a "seed", a technique akin to the use of a  seed crystal  in  crystallization . [ citation needed ]


The  pylorus  ( / p aɪ ˈ l ɔːr ə s /  or  / p ɪ ˈ l oʊ r ə s / ) connects the  stomach  to the  duodenum . The pylorus is considered as having two parts, the  pyloric antrum  (opening to the body of the stomach) and the  pyloric canal  (opening to the duodenum). The  pyloric canal  ends as the  pyloric orifice , which marks the junction between the stomach and the duodenum. The orifice is surrounded by a  sphincter , a band of muscle, called the  pyloric sphincter .
The word  pylorus  comes from  Greek  πυλωρός, via  Latin . The word  pylorus  in Greek means "gatekeeper", related to "gate" ( Greek :  pyle ) and is thus linguistically related to the word " pylon ". [ 1 ]


The pylorus is the furthest part of the stomach that connects to the  duodenum . It is divided into two parts, the  antrum , which connects to the body of the stomach, and the  pyloric canal , which connects to the duodenum. [ 2 ]


The antrum  also called the  gastric antrum  or the  pyloric antrum  is the initial portion of the pyloric region. It is near the bottom of the stomach, proximal to the pyloric sphincter, which separates the stomach and the duodenum.  It may temporarily become partially or completely shut off from the remainder of the stomach during  digestion  by  peristaltic  contraction of the prepyloric  sphincter ; it is  demarcated , sometimes, from the pyloric canal by a slight groove.


The  pyloric canal  ( Latin :  canalis pyloricus ) is the opening between the stomach and the duodenum. [ 3 ]  The wall thickness of the pyloric canal is up to 3 millimeters (mm) in infants younger than 30 days, [ 4 ]  and up to 8 mm in adults. [ 5 ]


The  pyloric sphincter , or  valve , is a strong ring of smooth muscle at the end of the pyloric canal which lets food pass from the stomach to the duodenum. It controls the outflow of gastric contents into the duodenum. [ 6 ]  It receives  sympathetic  innervation from the  celiac ganglion .


Under  microscopy , the pylorus contains numerous  glands , including  gastric pits , which constitute about half the depth of the pyloric  mucosa .  They consist of two or three short closed tubes opening into a common duct or mouth. These tubes are wavy, and are about one-half the length of the duct. The duct is lined by  columnar cells , continuous with the epithelium lining the surface of the  mucous membrane  of the stomach, the tubes by shorter and more cubical cell which are finely granular. The glands contain mucous cells and  G cells  that secrete  gastrin . [ 7 ]


The pylorus also contains scattered  parietal cells  and  neuroendocrine cells . These  endocrine  cells include  D cells , which release  somatostatin , [ 8 ]  responsible for shutting off acid secretion. (There is a second hormone-sensitive population near the  fundus .)
Unstriated muscles, which are entirely involuntary, are located at the pylorus.


The pylorus is one component of the  gastrointestinal system . Food from the  stomach , as  chyme , passes through the pylorus to the  duodenum . The pylorus, through the pyloric sphincter, regulates entry of food from the stomach into the duodenum.


In such conditions as  stomach cancer , tumours may partly block the pyloric canal. A special tube can be implanted surgically to connect the stomach to the duodenum so as to facilitate the passage of food from one to the other. The surgery to place this tube is called a  gastroduodenostomy .


Pyloric stenosis  refers to a pylorus that is narrow. This is due to congenital  hypertrophy  of the pyloric sphincter. The lumen of the pylorus is narrower, and less food is able to pass through. This problem is often detected in the early weeks of life. When it is present, a newborn baby may projectile vomit after eating, but despite vomiting remain hungry. Pyloric stenosis may be managed by the insertion of a  stent , or through surgical cutting of the pyloric sphincter, a  pyloromyotomy . [ 9 ]


The  rectum  ( pl. :  rectums  or  recta ) is the final straight portion of the  large intestine  in humans and some other  mammals , and the  gut  in others. Before expulsion through the  anus  or  cloaca , the rectum stores the  feces  temporarily. The adult human rectum is about 12 centimetres (4.7 in) long, [ 2 ]  and begins at the rectosigmoid junction (the end of the  sigmoid colon ) at the level of the third sacral vertebra or the sacral promontory depending upon what definition is used. [ 3 ]  Its diameter is similar to that of the sigmoid colon at its commencement, but it is dilated near its termination, forming the  rectal ampulla . [ 4 ]  It terminates at the level of the anorectal ring (the level of the  puborectalis  sling) or the  dentate line , again depending upon which definition is used. [ 3 ]  In humans, the rectum is followed by the  anal canal , which is about 4 centimetres (1.6 in) long, before the gastrointestinal tract terminates at the  anal verge . The word rectum comes from the  Latin   rēctum   intestīnum , meaning  straight intestine .


The human rectum is a part of the  lower gastrointestinal tract . The rectum is a continuation of the  sigmoid colon , and connects to the  anus . The rectum follows the shape of the  sacrum  and ends in an expanded section called an ampulla where  feces  is stored before its release via the  anal canal . An ampulla (from  Latin    bottle ) is a cavity, or the dilated end of a duct, shaped like a Roman  ampulla . [ 5 ]  The rectum joins with the sigmoid colon at the level of  S3 , and joins with the anal canal as it passes through the  pelvic floor  muscles. [ 5 ]


Unlike other portions of the colon, the rectum does not have distinct  taeniae coli . [ 6 ]  The taeniae blend with one another in the sigmoid colon five centimeters above the rectum, becoming a singular longitudinal muscle that surrounds the rectum on all sides for its entire length. [ 7 ] [ 6 ]


The blood supply of the rectum changes between the top and bottom portions. [ 8 ]  The top two thirds is supplied by the  superior rectal artery . The lower third is supplied by the  middle  and  inferior rectal arteries . [ 8 ]


The superior rectal artery is a single artery that is a continuation of the  inferior mesenteric artery , when it crosses the  pelvic brim . [ 8 ]  It enters the mesorectum at the level of S3, and then splits into two branches, which run at the lateral back part of the rectum, and then the sides of the rectum. These then end in branches in the submucosa, which join with ( anastamose ) with branches of the middle and inferior rectal arteries. [ 8 ]


The microanatomy of the wall of the rectum is similar to the rest of the  gastrointestinal tract ; [ 9 ]  namely, that it possesses a mucosa with a lining of  a single layer of column-shaped cells  with mucus-secreting  goblet cells  interspersed, resting on a  lamina propria , with a layer of smooth muscle called  muscularis mucosa . This sits on an underlying  submucosa  of connective tissue, surrounded by a  muscularis propria  of two bands of muscle, an inner circular band and an outer longitudinal one. [ 10 ]  There are a higher concentration of goblet cells in the rectal mucosa than other parts of the gastrointestinal tract. [ 9 ]


The lining of the rectum changes sharply at the line where the rectum meets the  anus . Here, the lining changes from the column-shaped cells of the rectum to  multiple layers of flat cells . [ 9 ]


The rectum acts as a temporary storage site for feces. The rectum receives fecal material from the  descending colon , transmitted through regular muscle contractions called  peristalsis . [ 11 ]  As the rectal walls expand due to the materials filling it from within, stretch receptors from the  nervous system  located in the rectal walls stimulate the desire to pass feces, a process called  defecation . [ 11 ]


An  internal  and  external anal sphincter , and resting contraction of the  puborectalis , prevent leakage of feces ( fecal incontinence ). As the rectum becomes more distended, the sphincters relax and a  reflex  expulsion of the contents of the rectum occurs. Expulsion occurs through contractions of the muscles of the rectum. [ 11 ]


The urge to voluntarily defecate occurs after the rectal pressure increases to beyond 18 mmHg; and reflex expulsion at 55 mmHg. In voluntary defecation, in addition to contraction of the rectal muscles and relaxation of the external anal sphincter, abdominal muscle contraction, and relaxation of the puborectalis muscle occurs. This acts to make the angle between the rectum and anus straighter, and facilitate defecation. [ 11 ]


For the diagnosis of certain ailments, a  rectal exam  may be done. These include  faecal impaction ,  prostatic cancer  and  benign prostatic hypertrophy  in men,  faecal incontinence , and internal  haemorrhoids . [ 12 ]  Forms of  medical imaging  used to examine the rectum include  CT scans  and MRI scans. An  ultrasound  probe may be inserted into the rectum to view nearby structures such as the prostate.


Colonoscopy  and  sigmoidoscopy  are forms of  endoscopy  that use a guided camera to directly view the rectum. The instruments may have the ability to take  biopsies  if needed, for diagnosis of diseases such as  cancer . A  proctoscope  is another instrument that is used to visualise the rectum.


Body temperature  can also be taken in the rectum. Rectal temperature can be taken by inserting a  medical thermometer  not more than 25 mm (0.98 in) into the rectum via the  anus . A  mercury thermometer  should be inserted for 3 to 5 minutes; a digital thermometer should remain inserted until it beeps. Normal rectal temperature generally ranges from 36 to 38 °C (97 to 100 °F) and is about 0.5 °C (32.9 °F) above oral (mouth) temperature and about 1 °C (34 °F) above  axilla  (armpit) temperature. [ citation needed ]  Availability of less invasive temperature-taking methods including tympanic (ear) and forehead thermometers has facilitated reduced use of this method.


Some medications are also administered via the rectum ( Latin :  per rectum ). [ 13 ]  By their definitions, suppositories are inserted, and enemas are injected into the rectum. [ 14 ] [ 15 ]  Medications might be given via the rectum to relieve constipation, to treat conditions near the rectum, such as fissures or haemorrhoids, or to give medications that are systemically active when taking them by mouth is not possible. [ 16 ]  People do not tend to like medications administered by this route because of both cultural issues, discomfort, and issues that may affect the medication working, such as leakage. [ 16 ]


One cause of  constipation  is  faecal impaction  in the rectum, in which a dry, hard  stool  forms. [ citation needed ]  Constipation is most commonly due to dietary and lifestyle factors such as inadequate  hydration , immobility, and lack of dietary fibre, although there are many potential causes. [ 17 ]  Such causes may include obstruction because of narrowing, local disease (such as Crohn's disease, fissures or haemorrhoids), or diseases affecting the neurological control of the bowel, or slow bowel transit time, including  spinal cord injury  and  multiple sclerosis ; use of medications such as  opioids , and conditions such as  diabetes mellitus , as well as severe illness. [ 17 ]  High calcium levels and  low thyroid activity  may also cause constipation. [ 17 ]


Testing may be carried out to investigate the cause. This may include  blood tests  such as  biochemistry ,  calcium  levels,  thyroid function tests . [ 17 ]  A digital rectal examination may be performed to see if there is stool in the rectum, and whether there is an obstruction. [ 17 ]  When symptoms such as weight loss, bleeding through the rectum, or pain are present, additional investigations such as a  CT scan  may be ordered. [ 17 ]  If constipation persists despite simple treatments, testing may also include  anal manometry  to measure pressures in the anus and rectum, electrophysiological studies, and magnetic resonance proctography. [ 17 ]


In general however, constipation is treated by improving factors such as hydration, exercise, and dietary fibre. [ 17 ]   Laxatives  may be used. Constipation that persists may require enemas or suppositories. Sometimes, use of the fingers or hand (manual evacuation) is required. [ citation needed ]  Although  peristalsis  in the colon delivers material to the rectum, laxatives such as  bisacodyl  or  senna  that induce peristalsis in the large bowel do not appear to initiate peristalsis in the rectum. They induce a sensation of rectal fullness and contraction that frequently leads to defecation, but without the distinct waves of activity characteristic of peristalsis. [ 18 ]


Other diseases of the rectum include:


Due to the proximity of the anterior wall of the rectum to the  vagina  in females or to the  prostate  in males, and the shared nerves thereof, the rectum is an  erogenous zone  and its  stimulation  or penetration can result in  sexual arousal . [ 19 ]


English  rectum  is derived from the Latin  intestinum rectum [ 20 ]  'straight gut', [ 21 ] [ 22 ]  a  calque [ 23 ] [ 24 ]  of  Ancient Greek  ἀπευθυσμένον ἔντερον, derived from ἀπευθύνειν,  to make straight , [ 25 ]  and ἔντερον,  gut , [ 25 ]  attested in the writings of Greek  physician   Galen . [ 23 ] [ 24 ]  During his anatomic investigations on animal corpses, Galen observed the rectum to be straight instead of curved as in humans. [ 23 ] [ 24 ]  The expressions ἀπευθυσμένον ἔντερον and intestinum rectum are therefore not appropriate descriptions of the rectum in humans.  Apeuthysmenon [ 26 ]   is the Latinization of ἀπευθυσμένον and  euthyenteron [ 27 ]  has a similar meaning  (εὐθύς 'straight [ 25 ] ). Much of the knowledge of the anatomy of the rectum comes from detailed descriptions provided by  Andreas Vesalius  in 1543. [ 28 ]


The  reticulum  is the second chamber in the four-chamber  alimentary canal  of a  ruminant  mammal.  Anatomically it is the smaller portion of the  reticulorumen  along with the  rumen . Together these two compartments make up 84% of the volume of the total stomach.


The reticulum is colloquially referred to as the  honeycomb ,  bonnet' , [ 1 ]  or  kings-hood . [ 1 ]   When cleaned and used for food, it is called " tripe ".


Heavy or dense feed and foreign objects will settle here. It is the site of  hardware disease  in cattle and because of the proximity to the heart this disease can be life-threatening.


The internal mucosa has a honeycomb shape. When looking at the reticulum with ultrasonography it is a crescent-shaped structure with a smooth contour. [ 2 ]  The reticulum is adjacent to the diaphragm, lungs, abomasum, rumen and liver. The heights of the reticular crests and depth of the structures vary across ruminant animal species. [ 3 ]   Grazing  ruminants have higher crests than  browsers . However, general reticulum size is fairly constant across ruminants of differing body size and feeding type.


Why nature selected the honeycomb shape as the internal mucosal structure of the reticulum. The main function of the reticulum is to store higher-density particles and generate biphasic contraction for the separation of food particles governed by the large amount of water in the reticulum. In The internal anatomical structure of the reticulum, the Honeycomb resembles the classical organic benzene structure; it is the most favourite structure of the nature. A hexagon has six sides, which allows it to distribute forces evenly across its structure, making it more stable and resistant to deformation. Additionally, the angles of a hexagon are evenly spaced, which helps to evenly distribute stress and prevent weak points, it has the angle that releases the most tension i.e.;120 degrees. The hexagon uses the least amount of material to hold the most weight that’s why higher density particles remain stay in the reticulum. High density particles may settle into the honeycomb structures and can be found after death. It is during the contractions of the reticulum that sharp objects can penetrate the wall and make their way to the heart. The hexagon is considered one of the strongest shapes in nature due to its ability to distribute force evenly and efficient. Hexagonal structures allow water molecules (with their one atom of oxygen and two of hydrogen) to group up together in the most efficient way and help in food particles separation. [ 4 ]


In a mature cow, the reticulum can hold around 5 gallons of liquid. The rumen and reticulum are very close in structure and function and can be considered as one organ. They are separated only by a muscular fold of tissue.


In immature ruminants, a reticular groove is formed by the muscular fold of the reticulum. This allows milk to pass by the reticulorumen straight into the abomasum.


The fluid contents of the reticulum play a role in particle separation. This is true both in domestic and wild ruminants. The separation takes place through biphasic contractions. In the first contraction, there is sending large particles back into the rumen while the reticulo-omasal orifice allows the passage of finer particles. In the second contraction, the reticulum contracts completely so the empty reticulum can refill with contents from the rumen. These contents are then sorted in the next biphasic contraction. [ 3 ]  The contractions occur in regular intervals. High density particles may settle into the honeycomb structures and can be found after death. It is during the contractions of the reticulum that sharp objects can penetrate the wall and make their way to the heart. Some ruminants, such as goats, also have monophasic contractions in addition to the biphasic contractions.


Retroperistalsis  is the reverse of the involuntary  smooth muscle  contractions of  peristalsis . It usually occurs as a precursor to  vomiting . Local irritation of the stomach, such as bacteria or food poisoning, activates the  emetic center  of the brain which in turn signals an imminent  vomiting reflex . Retroperistalsis begins in the small intestine and  pyloric sphincter . [ 1 ]  Food then moves in the opposite direction, often from the  duodenum  into the  stomach .


Retroperistalsis occurs pathologically during vomiting and physiologically at the first part of the duodenum where it protects from high acidity of food, and also at the  terminal ileum , where an amount of water and  electrolytes  are absorbed to assist  defecation . [ citation needed ]


The  rumen , also known as a  paunch , is the largest stomach compartment in  ruminants .  [ 1 ]  The rumen and the reticulum make up the  reticulorumen  in  ruminant  animals. [ 2 ]  The diverse  microbial  communities in the rumen allows it to serve as the primary site for microbial  fermentation  of ingested feed, which is often fiber-rich roughage typically indigestible by  mammalian  digestive systems. [ 2 ] [ 3 ]  The rumen is known for containing unique microbial networks within its multiple sac compartments to break down nutrients into usable energy and fatty acids.  [ 4 ]


The rumen is composed of five muscular sacs: cranial sac, ventral sac, dorsal sac, caudodorsal sac, and caudoventral blind sac. Each of these areas contain unique microbial communities, environments, and physical abilities that influence digestion.  [ 1 ] [ 4 ]


The outer lining of the rumen, known as the epithelium, serves as a protective layer and contributes to the metabolic processing of  fermentation  products.  [ 3 ]


The inner lining of the rumen wall is covered in small fingerlike projections called papillae, which aid in nutrient absorption. [ 1 ]  The  reticulum  is lined with ridges that form a  hexagonal   honeycomb  pattern. [ 1 ]  These features increase the surface area of the  reticulorumen  wall, facilitating the absorption of  volatile fatty acids  and capture of smaller digesta particles. [ 1 ]


The rumen and the  reticulum  differ with regard to the makeup of the lining but account for approximately 80% of total  ruminant  stomach volume.  [ 1 ]


Digestion in the rumen and  reticulorumen  occurs through  fermentation  by diverse  microbe  communities to optimize resources from nutrient dense feed. [ 5 ]  Millions of  microorganisms , including  bacteria ,  archaea ,  viruses ,  fungi , and  protozoa , are known to reside in the  reticulorumen  and are essential to digest structural  carbohydrates , like  lignocellulose  ( hemicellulose  and  cellulose ), non-structural  carbohydrates  ( starch ,  sugar , and  pectin ),  lipids , and  nitrogenous compounds  ( proteins ,  peptides , and  amino acids ). [ 5 ]


Both non-structural and structural carbohydrates are  hydrolysed  to  monosaccharides  or  disaccharides  by microbial  enzymes . The resulting  mono - and  disaccharides  are transported into the microbes.  Once within microbial cell walls, the  mono - and  disaccharides   may be assimilated into microbial biomass or fermented to  volatile fatty acids  (VFAs), such as  acetate ,  propionate ,  butyrate ,  lactate ,  valerate  and other  branched-chain  VFAs via  glycolysis  and other biochemical pathways to yield energy for the microbial cell. [ 1 ] [ 3 ]   Most VFAs are absorbed across the  reticulorumen  wall, directly into the bloodstream, and are used by the ruminant as substrates for energy production and  biosynthesis . [ 1 ] [ 6 ]  Some  branched chain VFAs  are incorporated into the  lipid membrane  of rumen microbes. VFAs provide large amounts of energy for ruminants and are critical to the health of the rumen and its microbiome. [ 7 ]


Lipids ,  lignin ,  minerals , and  vitamins  play a less prominent role in digestion than carbohydrates and protein, but they are still critical in many ways.  Lipids  are partly hydrolysed and hydrogenated, and  glycerol , if present in the lipid, is fermented. Lipids are otherwise inert in the rumen. Some carbon from carbohydrate or protein may be used for  de novo synthesis  of microbial lipid.  High levels of lipid, particularly unsaturated lipid, in the rumen are thought to poison microbes and suppress fermentation activity.  Lignin , a phenolic compound, is recalcitrant to digestion, through it can be solubilized by fungi. Lignin is thought to shield associated nutrients from digestion and hence limits degradation. Minerals are absorbed by microbes and are necessary to their growth.  Microbes in turn synthesize many vitamins, such as  cyanocobalamin , in great quantities—often great enough to sustain the ruminant even when vitamins are highly deficient in the diet.


The protein ingested is either  degradable intake protein  or  undegradable intake protein , or  rumen bypass protein .  [ 1 ]  Protein is hydrolysed to  peptides  and  amino acids  by microbial enzymes, which are subsequently transported across the microbial cell wall for assimilation into cell biomass, primarily.  Peptides, amino acids, ammonia, and other sources of nitrogen originally present in the feed can also be used directly by microbes with little to no hydrolysis. In situations in which nitrogen for microbial growth is in excess, protein and its derivatives can also be fermented to produce energy, yielding  ammonia . Excess  ammonia  is absorbed by the rumen and converted into  urea  in the liver. Non-amino acid nitrogen is used for synthesis of microbial amino acids. [ 1 ]


Ruminants  have access to food-sourced protein and microbial proteins produced by the  microbes  in the rumen. [ 1 ]  This creates a  symbiotic  relationship between the  ruminant  and the  microbial  communities, as the  microbes  can be used as a protein source when washed into the  abomasum  section of the digestive tract. [ 1 ]


Digested food (digesta) in the rumen is not uniform, but rather stratified into gas, liquid, and particles of different sizes, densities, and other physical characteristics. Additionally, the digesta is subject to extensive mixing and complicated flow paths upon entry into the rumen. Though they may seem trivial at first, these complicated stratification, mixing, and flow patterns of digesta are a key aspect of digestive activity in the ruminant and thus warrant detailed discussion.


After being swallowed, food travels down the  oesophagus  and is deposited in the  dorsal  part of the reticulum. Contractions of the reticulorumen propel and mix the recently ingested feed into the ruminal mat.  The mat is a thick mass of digesta, consisting of partially degraded, long, fibrous material.  Most material in the mat has been recently ingested, and as such, has considerable fermentable substrate remaining.  Microbial fermentation proceeds rapidly in the mat, releasing many gases.  Some of these gases are trapped in the mat, causing the mat to be buoyant.  As fermentation proceeds, fermentable substrate is exhausted, gas production decreases, and particles lose buoyancy due to loss of entrapped gas.  Digesta in the mat hence goes through a phase of increasing buoyancy followed by decreasing buoyancy.  Simultaneously, the size of digesta particles–relatively large when ingested–is reduced by microbial fermentation and, later, rumination. Incomplete digestion of plant material here will result in the formation of a type of  bezoar  called Phytobezoars. At a certain point, particles are dense and small enough that they may “fall” through the rumen mat into the ventral sac below, or they may be swept out of the rumen mat into the reticulum by liquid gushing through the mat during ruminal contractions. Once in the ventral sac, digesta continues to ferment at decreased rates, further losing buoyancy and decreasing in particle size.  It is soon swept into the ventral reticulum by ruminal contractions.


In the ventral reticulum, less dense, larger digesta particles may be propelled up into the oesophagus and mouth during contractions of the reticulum.  Digesta is chewed in the mouth in a process known as  rumination , then expelled back down the oesophagus and deposited in the dorsal sac of the reticulum, to be lodged and mixed into the ruminal mat again.  Denser, small particles stay in the ventral reticulum during reticular contraction, and then during the next contraction may be swept out of the reticulorumen with liquid through the  reticulo-omasal orifice , which leads to the next chamber in the ruminant animal's alimentary canal, the  omasum .


Water and saliva enter through the rumen to form a liquid pool.  Liquid will ultimately escape from the reticulorumen from absorption through the wall, or through passing through the reticulo-omasal orifice, as digesta does.  However, since liquid cannot be trapped in the mat as digesta can, liquid passes through the rumen much more quickly than digesta does.  Liquid often acts as a carrier for very small digesta particles, such that the dynamics of small particles is similar to that of liquid.


The uppermost area of the rumen, the headspace, is filled with  gases  (such as  methane ,  carbon dioxide , and, to a much lower degree,  hydrogen ) released from  fermentation  and  anaerobic respiration  of food. These gases are regularly expelled from the reticulorumen through the mouth, in a process called  eructation .


The different sacs of the rumen allow for varying ecological niches for microbes in the reticulorumen, including  bacteria ,  protozoa ,  fungi ,  archaea , and  viruses . [ 7 ]  Each microbial community depends on a variety of enzymes to breakdown  lignocellulose , nonstructural  carbohydrates ,  nitrogenous compounds , and  lipids . [ 5 ]


Bacteria, along with  protozoa , are the predominant microbes and by mass account for 40-60% of total microbial matter in the rumen. [ 8 ] [ 9 ]  They are categorized into several functional groups, such as  fibrolytic ,  amylolytic , and  proteolytic  types, which preferentially digest structural carbohydrates, non-structural carbohydrates, and protein, respectively.  Protozoa  (40-60% of microbial mass) derive their nutrients through  phagocytosis  of other microbes, and degrade and digest feed  carbohydrates , especially starch and sugars, and protein. [ 8 ]


Ruminal  fungi  make up 5-10% of microbes and are absent on diets poor in fibre. [ 9 ]   Fungi occupy an important niche in the rumen because they hydrolyse some ester linkages between  lignin  and  hemicellulose  or  cellulose , and help break down digesta particles.   Archaea , approximately 3% of total microbes, are mostly  autotrophic  methanogens and produce  methane  through  anaerobic respiration . [ 9 ]   Most of the hydrogen produced by  bacteria ,  protozoa  and  fungi  is used by these  methanogens  to reduce  carbon dioxide  to  methane . [ 9 ]   Viruses  are present in unknown numbers and have not been well studied.  However, they can  lyse  microbes, releasing their contents for other microbes to assimilate and ferment in a process called  microbial recycling , although recycling through the predatory activities of protozoa is quantitatively more important. [ 9 ]


Microbes  in the  reticulorumen  eventually flow out into the  omasum  and the remainder of the  alimentary canal .  Under normal  fermentation  conditions the environment in the  reticulorumen  is weakly acidic and is populated by microbes that are adapted to a pH between roughly 5.5 and 6.5; since the abomasum is strongly acidic (pH 2 to 4), it acts as a barrier that largely kills  reticulorumen   flora  and  fauna  as they flow into it. [ 1 ]   Subsequently, microbial  biomass  is digested in the  small intestine  and smaller molecules (mainly  amino acids ) are absorbed and transported in the portal vein to the liver. [ 1 ]   The digestion of these  microbes  in the small intestine is a major source of nutrition, as microbes usually supply some 60 to 90% of the total amount of amino acids absorbed. On starch-poor diets, they also provide the predominant source of glucose absorbed from the small intestinal contents. [ 1 ] [ 8 ]


The feed contained within the reticulorumen, known as "paunch waste", has been studied as a fertiliser for use in  sustainable agriculture . [ 10 ]


At birth, the rumen  organ , rumen  epithelium , and rumen  microbiota  are not fully developed and are metabolically nonfunctional. [ 3 ]  The developing rumen does not display the level of keratinization seen in the mature organ. [ 3 ]   Generally, the most receptive time for rumen development is between the  postnatal  and  weaning  periods. Over this period, rumen organ and  epithelium  growth, along with the establishment of rumen  microbiota , will prove to be essential to rumen development. [ 11 ]  This process is influenced by the introduction of solid food and the establishment of  fermentation  in the rumen. [ 3 ]  Additionally, there must be an adequate amount of  short chain fatty acids , produced during  fermentation , to properly develop the papillae. [ 3 ]


Papillae  growth in rumen  epithelium  is essential for rumen functionality. Papillae increase the surface area inside of the rumen and allow for a considerable increase in nutrient  absorption  inside of the rumen. [ 1 ]  Distinguishing a developed from an undeveloped rumen is simplified by observing the carpeting of tissue surrounding the interior of the rumen, as an undeveloped rumen maintains a smooth, papillae-lacking outer surface, and a developed rumen possesses thick, papillae-full walls. [ 3 ] [ 12 ]


Due to ruminants being born with a sterile  gastrointestinal tract , the developing rumen must be exposed to an array of microflora at an early stage. Specific diets in which  microflora  promote an  anaerobic  environment suitable for fermentation in the rumen are favored. [ 11 ]  Furthermore, feeds must be tailored to the needs of the specific  ruminants , as developing  ruminants  who have been on a strict liquid feed diet will possess different  microflora  when compared to that of a developing  ruminant  fed with a combination of a dry and liquid feed. [ 3 ]  This is due to the nutrients ingested by the animal not entering into the rumen stomach compartment, as it is instead bypassed by the reflexive closure of the  esophageal  groove. [ 11 ]


The most abundant bacteria present in the rumen microbiome include  Prevotella ,  Butyrivibrio , and  Ruminococcus . [ 13 ]  This is due to ruminant organisms ingesting high-forage, commonly grass-based diets. Their typical high-forage diets cause this significant demand for cellulose digesting bacteria to be ever-present. Other bacteria, such as  Lachnospira multiparus, Prevotella ruminicola,  and  Butyrivibrio fibrisolvens , play essential roles in the creation of  volatile fatty acids  (VFAs). [ 13 ]  Specific feeds can stimulate this extensive bacterial growth in the rumen and therefore aid in the production of these volatile fatty acids, which play a major role in rumen epithelium growth, capillary development, and papillae formation. [ 14 ]  Previous research identified the significant impact of volatile fatty acids on rumen development through the effects of the inter-ruminal insertion of  acetate ,  propionate , and  butyrate . [ 15 ]  The most visually notable and impactful of these volatile fatty acids was butyrate, which is synthesized naturally in ruminants through multiple anaerobic fermentation pathways of dietary substrates. [ 16 ]  Butyrate, mainly expressed in epithelial tissue lining, is involved in regulating a plethora of ruminant epithelial cell genes. Generally, butyrate regulates gene expression by acting on cell cycle control pathways. [ 17 ]  In the epithelial wall of the rumen, butyrate regulates epithelial cell gene expression to increase blood flow and papilla proliferation. [ 15 ]


Developing feeds to support the microbiome growth of both production and pet ruminant animals is vital; both for the overall health of the maturing animal and for reducing the costs associated with raising that animal. In the production animal realm, feeding can account for up to 75% of the overall cost associated with that animal, making it crucial to identify and satisfy the nutritional demands of the rumen. [ 18 ]  Sampling microbial DNA from rumen epithelial cells has led to the identification of microbial genes and functional pathways associated with animal growth factors. [ 19 ]  Microbial clusters in the rumen possess genes associated with many animal growth-related factors. Protein encoding genes that encode for bacterial cell functions, such as  aguA, ptb , K01188, and  murD , also are associated with the animal’s average daily weight gain. [ 18 ]  Furthermore, vitamin B12 related genes, including  cobD, tolC, and fliN,  are also related to the daily feed intake of the animal. [ 18 ]


S cells  are cells which release  secretin , found in the  jejunum  and  duodenum . [ 1 ]  They are stimulated by a drop in  pH  to 4 or below in the  small intestine 's lumen. The released secretin will increase the secretion of  bicarbonate  (HCO 3 − ) into the lumen, via the  pancreas . This is primarily accomplished by an increase in  cyclic AMP  that activates  CFTR  to release chloride anions into the lumen. The luminal Cl −  is then involved in a  bicarbonate transporter protein  exchange, in which the chloride is reabsorbed by the cell and HCO 3 −  is secreted into the  lumen . S cells are also one of the main producers of  cyclosamatin .


Saccharomyces cerevisiae  ( / ˌ s ɛr ə ˈ v ɪ s i . iː / ) ( brewer's yeast  or  baker's yeast ) is a species of  yeast  (single-celled  fungal  microorganisms). The species has been instrumental in  winemaking ,  baking , and  brewing  since ancient times. It is believed to have been originally isolated from the skin of  grapes . [ a ]  It is one of the most intensively studied  eukaryotic   model organisms  in  molecular  and  cell biology , much like  Escherichia coli  as the model  bacterium . It is the microorganism which causes many common types of  fermentation .  S. cerevisiae  cells are round to ovoid, 5–10  μm  in diameter. It reproduces by  budding . [ 1 ]


Many  proteins  important in human biology were first discovered by studying their  homologs  in yeast; these proteins include  cell cycle  proteins,  signaling proteins , and protein-processing  enzymes .  S. cerevisiae  is currently the only yeast cell known to have  Berkeley bodies  present, which are involved in particular secretory pathways.  Antibodies  against  S. cerevisiae  are found in 60–70% of patients with  Crohn's disease  and 10–15% of patients with  ulcerative colitis , and may be useful as part of a panel of serological markers in differentiating between inflammatory bowel diseases (e.g. between ulcerative colitis and Crohn's disease), their localization and severity. [ 2 ]


" Saccharomyces " derives from  Latinized   Greek  and means "sugar-mould" or "sugar-fungus",  saccharon  (σάκχαρον) being the combining form "sugar" and  myces  (μύκης) being " fungus ". [ 3 ] [ 4 ]   cerevisiae  comes from Latin and means "of beer". [ 5 ]   Other names for the organism are:


This species is also the main source of  nutritional yeast  and  yeast extract . [ citation needed ]


In the 19th century, bread bakers obtained their yeast from beer brewers, and this led to sweet-fermented breads such as the Imperial " Kaisersemmel " roll, [ 7 ]
which in general lacked the sourness created by the acidification typical of  Lactobacillus . However, beer brewers slowly switched from top-fermenting ( S. cerevisiae ) to bottom-fermenting ( S. pastorianus ) yeast. The  Vienna Process  was developed in 1846. [ 8 ]
While the innovation is often popularly credited for using steam in baking ovens, leading to a different crust characteristic, it is notable for including procedures for high milling of grains (see Vienna grits [ 9 ] ),
cracking them incrementally instead of mashing them with one pass; as well as better processes for growing and harvesting top-fermenting yeasts, known as press-yeast. [ 10 ]


Refinements in microbiology following the work of  Louis Pasteur  led to more advanced methods of culturing pure strains. In 1879, Great Britain introduced specialized growing vats for the production of  S. cerevisiae , and in the United States around the turn of the 20th century centrifuges were used for concentrating the yeast, [ 11 ]
turning yeast production into a major industrial process which simplified its distribution, reduced unit costs and contributed to the commercialization and commoditization of bread and beer. Fresh "cake yeast" became the standard leaven for bread bakers in much of the Western world during the early 20th century. [ 12 ]


During  World War II ,  Fleischmann's  developed a  granulated  active dry yeast for the United States armed forces, which did not require refrigeration and had a longer shelf-life and better temperature tolerance than fresh yeast; it is still the standard yeast for US military recipes. The company created yeast that would rise twice as fast, cutting down on baking time.  Lesaffre  would later create instant yeast in the 1970s, which has gained considerable use and market share at the expense of both fresh and dry yeast in their various applications. [ citation needed ]


In nature, yeast cells are found primarily on ripe fruits such as grapes (before maturation, grapes are almost free of yeasts). [ 13 ]   S. cerevisiae  can also be found year-round in the bark of  oak trees . [ 14 ]  Since  S. cerevisiae  is not airborne, it requires a vector to move. [ 15 ]


Queens of social wasps overwintering as adults ( Vespa crabro  and  Polistes  spp.) can harbor yeast cells from autumn to spring and transmit them to their progeny. [ 16 ]  The intestine of  Polistes dominula , a social wasp, hosts  S. cerevisiae  strains as well as  S. cerevisiae  ×  S. paradoxus  hybrids. Stefanini et al. (2016) showed that the intestine of  Polistes dominula  favors the mating of  S. cerevisiae  strains, both among themselves and with  S. paradoxus  cells by providing environmental conditions prompting cell  sporulation  and spores germination. [ 17 ]


The optimum temperature for growth of  S. cerevisiae  is 30–35 °C (86–95 °F). [ 16 ]


Two forms of yeast cells can survive and grow:  haploid  and  diploid . The haploid cells undergo a simple  lifecycle  of  mitosis  and growth, and under conditions of high stress will, in general, die. This is the  asexual  form of the fungus. The diploid cells (the preferential 'form' of yeast) similarly undergo a simple lifecycle of mitosis and  growth . The rate at which the mitotic cell cycle progresses often differs substantially between haploid and diploid cells. [ 18 ]  Under conditions of  stress , diploid cells can undergo  sporulation , entering  meiosis  and producing four haploid  spores , which can subsequently mate. This is the  sexual  form of the  fungus . Under optimal conditions, yeast cells can double their population every 100 minutes. [ 19 ] [ 20 ]  However, growth rates vary enormously between strains and between environments. [ 21 ]  Mean  replicative  lifespan is about 26 cell divisions. [ 22 ] [ 23 ]


In the wild, recessive deleterious mutations accumulate during long periods of  asexual reproduction  of diploids, and are purged during  selfing : this purging has been termed "genome renewal". [ 24 ] [ 25 ]


All  strains  of  S. cerevisiae  can grow  aerobically  on  glucose ,  maltose , [ 26 ]  and  trehalose [ 27 ]  and fail to grow on  lactose  and  cellobiose . However, growth on other  sugars  is variable.  Galactose  and  fructose  are shown to be two of the best fermenting sugars. The ability of yeasts to use different sugars can differ depending on whether they are grown aerobically or anaerobically. Some strains cannot grow anaerobically on  sucrose  and trehalose. [ citation needed ]


All strains can use  ammonia  and  urea  as the sole  nitrogen  source, but cannot use  nitrate , since they lack the ability to reduce them to  ammonium   ions . They can also use most  amino acids , small  peptides , and nitrogen bases as nitrogen sources.  Histidine ,  glycine ,  cystine , and  lysine  are, however, not readily used.  S. cerevisiae  does not excrete  proteases , so extracellular protein cannot be metabolized.


Yeasts  also have a requirement for  phosphorus , which is assimilated as a dihydrogen phosphate ion, and  sulfur , which can be assimilated as a  sulfate  ion or as organic sulfur compounds such as the  amino acids  methionine and cysteine. Some metals, like  magnesium ,  iron ,  calcium , and  zinc , are also required for good growth of the yeast.


Concerning organic requirements, most strains of  S. cerevisiae  require  biotin . [ 28 ]  Indeed, a  S. cerevisiae -based growth assay laid the foundation for the isolation, crystallization, and later structural determination of biotin. Most strains also require  pantothenate  for full growth. In general,  S. cerevisiae  is prototrophic for vitamins.


Yeast has two mating types,  a  and α ( alpha ), which show primitive aspects of sex differentiation. [ 29 ]  As in many other eukaryotes, mating leads to  genetic recombination , i.e. production of novel combinations of chromosomes. Two  haploid  yeast cells of opposite mating type can mate to form  diploid  cells that can either  sporulate  to form another generation of haploid cells or continue to exist as diploid cells. Mating has been exploited by biologists as a tool to combine genes, plasmids, or proteins at will. [ citation needed ]


The mating pathway employs a  G protein-coupled receptor ,  G protein ,  RGS protein , and three-tiered  MAPK  signaling cascade that is homologous to those found in humans. This feature has been exploited by biologists to investigate basic mechanisms of  signal transduction  and  desensitization . [ citation needed ]


Growth in yeast is synchronized with the growth of the  bud , which reaches the size of the mature cell by the time it separates from the parent cell. In well nourished, rapidly growing yeast  cultures , all the cells have buds, since bud formation occupies the whole  cell cycle . Both mother and daughter cells can initiate bud formation before cell separation has occurred. In yeast cultures growing more slowly, cells lacking buds can be seen, and bud formation only occupies a part of the cell cycle. [ citation needed ]


Cytokinesis  enables budding yeast  Saccharomyces cerevisiae  to divide into two daughter cells.  S. cerevisiae  forms a bud which can grow throughout its cell cycle and later leaves its mother cell when mitosis has completed. [ 30 ]


S. cerevisiae  is relevant to cell cycle studies because it divides asymmetrically by using a polarized cell to make two daughters with different fates and sizes. Similarly,  stem cells  use asymmetric division for self-renewal and differentiation. [ 31 ]


For many cells, M phase does not happen until S phase is complete. However, for entry into mitosis in  S. cerevisiae  this is not true. Cytokinesis begins with the budding process in late G1 and is not completed until about halfway through the next cycle. The assembly of the spindle can happen before S phase has finished duplicating the chromosomes. [ 30 ]  Additionally, there is a lack of clearly defined G2 in between M and S. Thus, there is a lack of extensive regulation present in higher eukaryotes. [ 30 ]


When the daughter emerges, the daughter is two-thirds the size of the mother. [ 32 ]  Throughout the process, the mother displays little to no change in size. [ 33 ]  The RAM pathway is activated in the daughter cell immediately after cytokinesis is complete. This pathway makes sure that the daughter has separated properly. [ 32 ]


Two interdependent events drive cytokinesis in  S. cerevisiae . The first event is contractile  actomyosin ring  (AMR) constriction and the second event is formation of the primary  septum  (PS), a chitinous cell wall structure that can only be formed during cytokinesis. The PS resembles in animals the process of extracellular matrix remodeling. [ 32 ]  When the AMR constricts, the PS begins to grow. Disrupting AMR misorients the PS, suggesting that both have a dependent role. Additionally, disrupting the PS also leads to disruptions in the AMR, suggesting both the actomyosin ring and primary septum have an interdependent relationship. [ 34 ] [ 33 ]


The AMR, which is attached to the cell membrane facing the cytosol, consists of actin and myosin II molecules that coordinate the cells to split. [ 30 ]  The ring is thought to play an important role in ingression of the plasma membrane as a contractile force. [ citation needed ]


Proper coordination and correct positional assembly of the contractile ring depends on septins, which is the precursor to the septum ring. These GTPases assemble complexes with other proteins. The septins form a ring at the site where the bud will be created during late G1. They help promote the formation of the actin-myosin ring, although this mechanism is unknown. It is suggested they help provide structural support for other necessary cytokinesis processes. [ 30 ]  After a bud emerges, the septin ring forms an hourglass. The septin hourglass and the myosin ring together are the beginning of the future division site. [ 35 ]


The septin and AMR complex progress to form the primary septum consisting of glucans and other chitinous molecules sent by vesicles from the Golgi body. [ 36 ]  After AMR constriction is complete, two secondary septums are formed by glucans. How the AMR ring dissembles remains poorly unknown. [ 31 ]


Microtubules do not play as significant a role in cytokinesis compared to the AMR and septum. Disruption of microtubules did not significantly impair polarized growth. [ 37 ]  Thus, the AMR and septum formation are the major drivers of cytokinesis. [ citation needed ]


When researchers look for an organism to use in their studies, they look for several traits. Among these are size, [ clarification needed ]  short generation time, accessibility [ clarification needed ] , ease of manipulation, genetics, [ clarification needed ]  conservation of mechanisms, [ clarification needed ]  and potential economic benefit. [ citation needed ]  The yeast species  Schizosaccharomyces pombe  and  S. cerevisiae  are both well studied; these two species diverged approximately  600 to 300  million years ago , and are significant tools in the study of  DNA damage  and  repair mechanisms . [ 39 ]


S. cerevisiae  has developed as a  model organism  because it scores favorably on a number of criteria.


For more than five decades  S. cerevisiae  has been studied as a model organism to better understand aging and has contributed to the identification of more mammalian genes affecting aging than any other model organism. [ 41 ]  Some of the topics studied using yeast are  calorie restriction , as well as in genes and cellular pathways involved in  senescence . The two most common methods of measuring aging in yeast are Replicative Life Span (RLS), which measures the number of times a cell divides, and Chronological Life Span (CLS), which measures how long a cell can survive in a non-dividing stasis state. [ 41 ]  Limiting the amount of glucose or amino acids in the  growth medium  has been shown to increase RLS and CLS in yeast as well as other organisms. [ 42 ]  At first, this was thought to increase RLS by up-regulating the sir2 enzyme; however, it was later discovered that this effect is independent of  sir2 . Over-expression of the genes sir2 and fob1 has been shown to increase RLS by preventing the accumulation of  extrachromosomal rDNA circles , which are thought to be one of the causes of senescence in yeast. [ 42 ]  The effects of dietary restriction may be the result of a decreased signaling in the TOR cellular pathway. [ 41 ]  This pathway modulates the cell's response to nutrients, and mutations that decrease TOR activity were found to increase CLS and RLS. [ 41 ] [ 42 ]  This has also been shown to be the case in other animals. [ 41 ] [ 42 ]  A yeast mutant lacking the genes  Sch9  and  Ras2  has recently been shown to have a tenfold increase in chronological lifespan under conditions of calorie restriction and is the largest increase achieved in any organism. [ 43 ] [ 44 ]


Mother cells give rise to progeny buds by mitotic divisions, but undergo replicative  aging  over successive generations and ultimately die. However, when a mother cell undergoes  meiosis  and  gametogenesis ,  lifespan  is reset. [ 45 ]  The replicative potential of  gametes  ( spores ) formed by aged cells is the same as gametes formed by young cells, indicating that age-associated damage is removed by meiosis from aged mother cells. This observation suggests that during meiosis removal of age-associated damages leads to  rejuvenation . However, the nature of these damages remains to be established.


During starvation of non-replicating  S. cerevisiae  cells,  reactive oxygen species  increase leading to the accumulation of  DNA damages  such as apurinic/apyrimidinic sites and double-strand breaks. [ 46 ]   Also in non-replicating cells the ability to  repair  endogenous double-strand breaks declines during chronological  aging . [ 47 ]


S. cerevisiae  reproduces by mitosis as diploid cells when nutrients are abundant. However, when starved, these cells undergo meiosis to form haploid spores. [ 48 ]


Evidence from studies of  S. cerevisiae  bear on the adaptive function of meiosis and  recombination .  Mutations  defective in genes essential for meiotic and mitotic recombination in  S. cerevisiae  cause increased sensitivity to  radiation  or  DNA damaging chemicals . [ 49 ] [ 50 ]  For instance, gene  rad52  is required for both meiotic recombination [ 51 ]  and mitotic recombination. [ 52 ]   Rad52  mutants have increased sensitivity to killing by  X-rays ,  Methyl methanesulfonate  and the DNA cross-linking agent  8-methoxypsoralen-plus-UVA , and show reduced meiotic recombination. [ 50 ] [ 51 ] [ 53 ]  These findings suggest that  recombination repair  during meiosis and mitosis is needed for repair of the different damages caused by these agents.


Ruderfer et al. [ 49 ]  (2006) analyzed the ancestry of natural  S. cerevisiae  strains and concluded that  outcrossing  occurs only about once every 50,000 cell divisions. Thus, it appears that in nature, mating is likely most often between closely related yeast cells. Mating occurs when haploid cells of  opposite mating type  MATa and MATα come into contact. Ruderfer et al. [ 49 ]  pointed out that such contacts are frequent between closely related yeast cells for two reasons. The first is that cells of opposite mating type are present together in the same  ascus , the sac that contains the cells directly produced by a single meiosis, and these cells can mate with each other. The second reason is that haploid cells of one mating type, upon cell division, often produce cells of the opposite mating type with which they can mate. The relative rarity in nature of meiotic events that result from  outcrossing  is inconsistent with the idea that production of  genetic variation  is the main selective force maintaining meiosis in this organism. However, this finding is consistent with the alternative idea that the main selective force maintaining meiosis is enhanced recombinational repair of DNA damage, [ 54 ]  since this benefit is realized during each meiosis, whether or not out-crossing occurs.


S. cerevisiae  was the first eukaryotic  genome  to be completely sequenced. [ 55 ]  The genome sequence was released to the  public domain  on April 24, 1996. Since then, regular updates have been maintained at the  Saccharomyces  Genome Database . This  database  is a highly annotated and cross-referenced database for yeast researchers. Another important  S. cerevisiae  database is maintained by the Munich Information Center for Protein Sequences (MIPS). Further information is located at the  Yeastract  curated repository. [ 56 ]


The  S. cerevisiae  genome is composed of about 12,156,677  base pairs  and 6,275  genes , compactly organized on 16 chromosomes. [ 55 ]  Only about 5,800 of these genes are believed to be functional. It is estimated at least 31% of yeast genes have  homologs  in the human genome. [ 57 ]  Yeast genes are classified using gene symbols (such as Sch9) or systematic names. In the latter case the 16 chromosomes of yeast are represented by the letters A to P, then the gene is further classified by a sequence number on the left or right arm of the chromosome, and a letter showing which of the two DNA strands contains its coding sequence. [ 58 ]


Examples:


The availability of the  S. cerevisiae  genome sequence and a set of deletion mutants covering 90% of the yeast genome [ 59 ]  has further enhanced the power of  S. cerevisiae  as a model for understanding the regulation of eukaryotic cells. A project underway to analyze the genetic interactions of all double-deletion mutants through  synthetic genetic array  analysis will take this research one step further. The goal is to form a functional map of the cell's processes.


As of 2010 [update]  a model of genetic interactions is most comprehensive yet to be constructed, containing "the interaction profiles for ~75% of all genes in the Budding yeast". [ 60 ]  This model was made from 5.4 million two-gene comparisons in which a double  gene knockout  for each combination of the genes studied was performed. The effect of the double knockout on the  fitness  of the cell was compared to the expected fitness. Expected fitness is determined from the sum of the results on fitness of single-gene knockouts for each compared gene. When there is a change in fitness from what is expected, the genes are presumed to interact with each other. This was tested by comparing the results to what was previously known. For example, the genes Par32, Ecm30, and Ubp15 had similar interaction profiles to genes involved in the Gap1-sorting module cellular process. Consistent with the results, these genes, when knocked out, disrupted that process, confirming that they are part of it. [ 60 ]


From this, 170,000 gene interactions were found and genes with similar interaction patterns were grouped together. Genes with similar genetic interaction profiles tend to be part of the same pathway or biological process. [ 61 ]  This information was used to construct a global network of gene interactions organized by function. This network can be used to predict the function of uncharacterized genes based on the functions of genes they are grouped with. [ 60 ]


Approaches that can be applied in many different fields of biological and medicinal science have been developed by yeast scientists. These include  yeast two-hybrid  for studying  protein interactions  and  tetrad analysis . Other resources, include a gene deletion library including ~4,700 viable haploid single gene deletion strains. A  GFP fusion strain library  used to study protein localisation and a  TAP tag library  used to purify protein from yeast cell extracts. [ citation needed ]


Stanford University's  yeast deletion project  created  knockout mutations  of every gene in the  S. cerevisiae  genome to determine their function. [ 62 ]


The yeast genome is highly accessible to manipulation, hence it is an excellent model for genome engineering.