This pathogen is easily identified in the stools using Gram stain and is responsible for cholera, a potentially fatal diarrheal disease in humans. Although cholera is now rare in developed countries, it remains a major cause of diarrheal morbidity and mortality in several developing countries. Indeed, with the occurrence of calamities, the spreading of cholera infection in overcrowded refugee camps is a potential significant threat everywhere. The infection is transmitted by the fecal–oral route and is spread through contaminated food and water, the period of incubation ranges from few hours to 5 days. The vast majority of infected subjects remain asymptomatic or experience a mild disease with watery stools, nausea or vomiting, and no significant dehydration. However, cholera may be severe in a number of subjects. Stools are classically described as “rice water” due to the presence of mucus in clear stools. Profuse watery diarrhea and vomiting lead to massive fluid and electrolyte losses that can occur at a rate of 1 l/h, causing a massive dehydration.

Virtually, all cases of non-O1 Vibrio infections in the USA are associated with eating raw shellfish and gastroenteritis ranges from a mild illness to profuse, watery diarrhea comparable to that seen in epidemic cholera. Diarrhea, abdominal cramps, and fever are the most common symptoms with nausea , vomiting, and bloody stools occurring less frequently. As with V. cholerae O1, the mainstay of therapy for diarrheal disease is oral rehydration [11, 12]. In cases of septicemia (which typically occurs in immunocompromised patients), supportive care and correction of shock are essential interventions associated with antibiotics (tetracycline). V. cholerae secretes several toxins, but the most important of these is cholera toxin (CT). CT consists in a single copy of the A subunit and five copies of the B subunit. B subunit binds to plasma membrane, while the A subunit activates adenylate cyclase resulting in elevated cAMP production. The production of cAMP activates the protein kinase A (PKA), which then phosphorylates the regulatory domain of CFTR [5, 13]. In addition to increased Cl⁻ secretion, absorption of Na⁺ is decreased through a cAMP-dependent mechanism in which the activity of both apical sodium transporters, NHE2 and NHE3, is decreased. This leads to an increase in NaCl levels in the intestinal lumen and drives water by osmotic force [14].

In addition to CT, V. cholerae produces other toxins that modulate ion secretion and alter barrier function to cause massive diarrhea. The toxins that directly affect ion secretion include the accessory CT (ACE), which stimulates Ca²⁺-dependent Cl⁻ secretion, the non-agglutinable Vibrio cholerae heat-stable enterotoxin (NAG-ST), which activates guanylyl cyclase, thus stimulating cGMP production, leading to protein kinase G (PKG)-mediated activation of CFTR and, finally, the V. cholerae cytolysin (VCC), which creates anion permeable pores into the cell wall [15].

Salmonella typhi and Salmonella paratyphi are Gram-negative, motile bacilli that colonize only humans. Therefore, the infection is acquired through close personal contact or through the ingestion of water or food contaminated with human feces. Typhoid fever continues to represent a global health problem, with more than 12.5 million cases/year [16]. After an incubation period ranging between 5 and 21 days, these bacteria cause a systemic illness characterized by fever, gastrointestinal symptoms, and occasionally neurological symptoms [17]. Chills, headache, cough, weakness, and muscle pain are frequent prodromes and most symptoms resolve within 4 weeks without antimicrobial treatment. However, Salmonella may spread and invade the bloodstream and extraintestinal districts causing a systemic disease defined as enteric fever. In this case, patients relapse with high fever, abdominal pain from inflammation of Peyer’s patches , and intestinal microperforation followed by secondary bacteremia with intestinal agents.

In contrast to S. typhi, infections with non-typhoidal salmonellae are increasing in developed countries. Patients at higher risk for infection include those with immunodeficiencies, age younger than 3 months, alterations in intestinal defenses (e.g., on those antacid treatment), impaired reticuloendothelial function (sickle cell and hemolytic anemia), and ingestion of antibiotics to which the organism is resistant. Reservoirs include a wide range of domestic and wild animals, including poultry, swine, cattle, rodents, and reptiles. Salmonella enteritidis is the leading reported cause of food—borne diarrheal outbreaks in the USA, with eggs and contaminated raw fruits and vegetables identified as major vehicles [18].

The incubation period is 6–45 h, after which fever, headache, vomiting, abdominal pain, and watery stools (which may contain blood, mucus, and leukocytes) appear, lasting from locus to few days. Severe extraintestinal infections can range from life-threatening sepsis to focal infections in the meninges, bones, and lungs. The microorganism is easily isolated from fresh stools or blood culture.

Clostridium difficile (CD), a Gram-positive anaerobe that forms spores (hence difficult to clear from hospital setting), is now recognized as the most common agent of nosocomial diarrhea [19]. In the past decade, a dramatic increase in the incidence of C. difficile infections (CDI) has been reported worldwide, possibly linked with an excess of antibiotics and other treatment responsible for microflora disruption but also linked with emergence of hypervirulent strains (e.g., NAP1/BI/027) and increased numbers of highly susceptible individuals.

Antibiotic treatment predisposes patients to C. difficile-associated disease in specific conditions [20]. The pathogenesis of C. difficile infection largely depends on the altered balance of the intestinal microbiota, allowing pathogenic strains of C. difficile to infect the intestine [21].

C. difficile causes a broad spectrum of intestinal diseases ranging from mild diarrhea to potentially fatal pseudomembranous colitis (PMC). C. difficile is a major agent of antibiotic-associated colitis [22] and produces three toxins, toxin A (TxA), toxin B (TxB), and a binary toxin (cytolethal distending toxin, CDT) [23]. Both TxA and TxB exert a cytotoxic effect in vitro [24, 25]. In particular, TxA and TxB induce the disaggregation of the actin cytoskeleton, cell rounding, cell death, and loss of intestinal epithelium barrier function leading to increased intestinal permeability and diarrhea [26]. CDT induces a redistribution of microtubules and the formation of long microtubule-based protrusions at the surface of intestinal epithelial cells [9] enhancing adherence and colonization of Clostridia [25, 27].

However, diarrhea is essentially due to a necroinflammatory reaction: C. difficile toxins trigger an extensive inflammatory cascade causing a rapidly progressive damage to host tissues resulting in fluid exudation [28]. In parallel, selected cytokines are also able to induce the activation of enteric nerves and induce Cl⁻ secretion. The onset of symptoms may begin from several days after antibiotic therapy is started until 2 months following cessation of treatment. Diarrhea and abdominal cramps are usually the presenting symptoms, followed by fever and chills in severe cases. Mild colitis, with bloody stools and mucus, particularly if they follow antibiotic treatment, should be considered potentially induced by C. difficile infection. Microbial culture, latex agglutination, tissue culture assay, and enzyme-linked immunosorbent assay (ELISA) are all used for the diagnosis of C. difficile infection.

However, the role of C.difficile in infancy and early childhood is still a matter of debate. This is an emerging agent of diarrhea whose role is limited or questionable in children below 36 months of age [29, 30]. It is also a major agent of antibiotic-induced diarrhea and of severe diarrhea in children with underlying chronic conditions such as inflammatory bowel diseases (IBD) [31].

Asymptomatic C. difficile colonization is common in infants (ranging from 2 to 75 % of healthy children) until 3 years of age and related to delivery mode and feeding [32]. Strains that usually are associated with diarrhea in adults may be detected in asymptomatic infants, and children may be a reservoir of pathogenic strains.

Many healthy children excrete C. difficile in their stools during early infancy, and rates as high as 45 % have been detected in infants attending day care nurseries and as many as 13 % were harboring toxigenic isolates [33].

Independently from the prevalence of carriers and the method to search for C. difficile, the incidence of C. difficile-related diseases in children has progressively increased in the last years, in parallel with data reported in adults, but with a relative more limited clinical impact. A recent 18-year cohort study reported a 12-fold increase of C. difficile infection incidence in children. Overall C. difficile infections increased from 2.6 (1991–1997) to 32.6 per 100,000 persons (2004–2009), with a prevalence of community acquired infection that increased from 2.2 to 23.4 per 100,000 in the same period [34]. However, a role of C. difficile as a severe pathogen in children at risk is supported by a solid data: For this reason, one potential alternative to standard therapy is the use of indigenous intestinal microorganisms from a healthy donor to restore the intestinal microbiota of infected patients [35, 36].

In children with chronic diseases such as IBD , C. difficile may act as severe opportunistic pathogen and trigger burst of the disease with a potentially severe course.

Shigella causes 250 million cases of diarrhea and 650,000 deaths worldwide per year. Shigella is a Gram negative, non-lactose fermenting, nonmotile bacillus. Shigella sonnei is the main type in industrialized countries, and S. flexneri and S. dysenteriae predominating in developing countries [37]. The latter is the most dangerous species as it produces Shiga toxin, which can lead to hemolytic uremic syndrome (HUS). Humans are the only natural hosts and transmission occurs by fecal–oral contact. The very low infective load (as few as ten organisms) makes Shigella highly contagious. Shigella causes a dysenteric diarrhea, and the cellular responses to various steps of the invasion process are the primary cause of inflammation. Shigella strains cross the epithelial barrier through the M-cells where they bind to basolateral TLR4 receptor. This causes the production of interleukin (IL)-6, IL-8, and the subsequent release of IL-1β, which attracts PMN cells [38]. Nuclear factor kappa B (NF-κB) and mitogen-activated protein kinase (MAPK)-signaling pathways are activated not only in Shigella-infected epithelial cells but also in uninfected bystander cells [39], and this cell–cell communication amplifies inflammation. However, Shigella extensively disrupts TJs. In polarized cultured T84 epithelial cells, ZO-1, claudin-1, and the phosphorylation status of occludin were all affected by S. flexneri [40]. There is a secondary mechanism that promotes diarrhea, triggered by serine protease autotransporters of Enterobacteriaceae (SPATES), three enterotoxins that alter transepithelial fluid movement and cytoskeletal structure [41]. After 1–4 days of incubation, shigellosis begins with fever, headache, malaise, anorexia, and occasional vomiting and watery diarrhea with progression to dysentery within hours to days. Unusual extraintestinal manifestations may occur, including HUS in children and thrombotic thrombocytopenic purpura in adults. Most episodes of shigellosis in otherwise healthy individuals resolve within 7 days. Shigellae are difficult to grow and are best isolated from fresh stools rapidly inoculated into selective culture plates incubated immediately at 37 °C.

These organisms are small, spiral-shaped Gram-negative bacilli that live in the intestines of both wild and domestic animals [42]. Common vehicles for human infection are poultry, unpasteurized milk, and contaminated water [43, 44]. After 3–6 days of incubation, symptoms abruptly begin. Campylobacter diarrhea may present as a typical gastroenteritis with vomiting and profuse diarrhea or as a colitis with abdominal pain and bloody stools, and abdominal pain that may mimic an appendicitis. Diarrhea usually lasts 4–5 days; the microorganism can be identified only from stool. Campylobacter vaccine development has proceeded cautiously, because of concerns about postexposure arthritis or Guillain–Barré syndrome. However, a monovalent, formalin-inactivated, C. jejuni whole-cell vaccine with a mucosal adjuvant has entered human trials.

Yersinia enterocolitica and Y. pseudotuberculosis are two important human enteropathogens widely distributed in the environment , with swine as the major reservoir. The incubation period is 3–7 days, with food-borne transmission. Yersinia’s preference for cool temperatures makes this pathogen more common in Northern Europe, Scandinavia, Canada, the USA, and Japan. Yersinia enterocolitis occurs more often in children younger than 5 years [45] and is characterized by fever, vomiting, exudative pharyngitis, cervical adenitis, abdominal pain, and watery diarrhea, which may contain blood [46, 47]. Diarrhea typically lasts for 14–22 days, but fecal excretion may persist for 7 weeks or longer. Abdominal complications include appendicitis , pseudoappendicitis, diffuse ulcerations of the intestine and colon, intestinal perforation, peritonitis , ileocecal intussusception , toxic megacolon, cholangitis, and mesenteric vein thrombosis. Bacteremic spread may result in abscess formation and granulomatous lesions in the liver, spleen, lungs, kidneys, and bone, as well as meningitis and septic arthritis. Yersinia infection can also be associated with extraintestinal sequelae including reactive arthritis, uveitis, Reiter’s syndrome, and erythema nodosum [48]. Yersinia may be isolated from stools or pharyngeal exudates on commonly used selective media, and appears as Gram-negative colonies after 2–14 days of growth at 25–28 °C.

E. coli is the most abundant facultative anaerobe of the human colonic flora and typically colonizes the gastrointestinal tract within few hours after birth. E. coli is a Gram-negative, lactose-fermenting motile bacillus of the family Enterobacteriaceae. Currently, 171 somatic (O) and 56 flagellar (H) antigens are recognized. E. coli includes a heterogeneous group of microorganisms capable to exert various possible interactions with the host, ranging from a role of mere harmless presence to that of a highly pathogenic organism [49]. Six distinct categories of E. coli are currently recognized as pathogens: enterotoxigenic E. coli (ETEC), enteropathogenic E. coli (EPEC), enterohemorrhagic E. coli (EHEC), diffusely adherent E. coli (DAEC), enteroaggregative E. coli (EAEC or EAggEC), and enteroinvasive E. coli (EIEC).

However, EPEC encode a T3SS and produces a characteristic attaching and effacing (A/E) lesion which is characterized by effacement of microvilli on the epithelial surface at the site of bacterial attachment. This typical lesion is responsible for the loss of absorptive surface and osmotic diarrhea [54]. Intestinal pathogens can disrupt the barrier by directly altering the distribution or phosphorylation status of TJ proteins . EPEC strains alter the phosphorylation status and distribution of TJ proteins, occludin and claudin-1, thereby disrupting the epithelial barrier function [10]. This increases paracellular permeability driving ion and water into the lumen upon electrochemical gradients ultimately causing diarrhea. The link between barrier disruption and diarrhea derives from studies involving the pro-inflammatory cytokine, tumor necrosis factor-α (TNFα) [55, 56]. These observations suggest that EPEC-induced diarrhea is a multifactorial process with alterations of electrolyte, solute, and water transport. EPEC causes a self-limited watery diarrhea with a short incubation period (6–48 h). This may be associated with fever, abdominal cramps, and vomiting, and EPEC is a leading cause of persistent diarrhea (lasting 14 days) in children in developing countries [57].

The microbiological diagnosis of EPEC-induced disease is performed with analytic methodologies different from those used by the standard microbiology laboratory, the most relevant being: (a) serotyping, (b) adherence assay, (c) FAS test, and (d) the specific detection of virulence-involved genes (bfpA and eae genes) using molecular biology techniques [58].

However, the cause–effect relationship between EPEC in the stools and diarrhea is hampered by the high number of healthy carriers.

EHEC adheres to epithelial cells, expresses a T3SS and causes A/E lesions much like EPEC. Unlike EPEC, infection with EHEC may cause severe symptoms including bloody diarrhea and life-threatening HUS. These symptoms are due to the production of Shiga toxin which elicits luminal fluid accumulation in the intestine [60, 61]. The predominant transmission is through the ingestion of contaminated food. Crampy abdominal pain and non-bloody diarrhea are the first symptoms, sometimes associated with vomiting. Diarrhea always becomes bloody and abdominal pain worsens, lasting 1–22 days. Fever is usually absent or low grade. In outbreaks, approximately 25 % of patients are hospitalized, 5–10 % develop HUS, and 1 % may have a poor outcome. The most dangerous complication of EHEC infection is HUS. This is usually diagnosed 2–14 days after the onset of diarrhea. Risk factors include young age, bloody diarrhea, fever, an elevated leukocyte count, and treatment with antimotility agents [62]. The most widely accepted indication for seeking E. coli O157:H7 infection is a patient with bloody diarrhea, in whom an accurate diagnosis may avoid unnecessary surgery.

Typical clinical features are a watery, mucoid, secretory diarrheal illness with low-grade fever, and little or no vomiting. However, bloody stools have been reported. Infection of EAggEG is detected by the isolation of E. coli from the stools of patients and the demonstration of the aggregative pattern in the HEp-2 assay. Establishing a cause–effect relationship is hampered by the high rate of asymptomatic colonization; if no other organism is implicated in the patients’ illness and EAggEC is isolated, the germ should be considered as a potential cause of diarrhea. A DNA-fragment probe has proven highly specific in the detection of EAggEC strains. Acute diarrhea is apparently self-limiting; however, more persistent cases may benefit from antibiotic and/or nutritional therapy, after susceptibility test [66].

Antibiotic therapy for acute bacterial gastroenteritis is not needed routinely but could be considered for specific pathogens or in defined clinical settings, including clinical condition and risk factors [68, 69].