**9. Microorganisms in biliary tract infections**

The majority of cases of acute cholecystitis are caused by an impacted gallstone blocking the gallbladder outlet, resulting in an increase in intraluminal pressure, gallbladder distension, and wall edema, and eventually gallbladder necrosis. During the early stages of acute cholecystitis, bile is normally sterile, and infection occurs as a side effect.

Biliary tract infection is a prevalent cause of bacteremia and is linked to a high rate of morbidity and mortality, especially in elderly individuals with comorbid conditions or when diagnosis and treatment are delayed. Enterobacteriaceae, which climb from the gastrointestinal system, are the most prevalent infectious organisms. Complications such as acute renal failure and septic shock are more likely in patients with bacteremia.

## **9.1 Bacterial causes of biliary tract infections**

The most frequently identified pathogens are Gram-negative microorganisms, primarily *Escherichia coli*, *Salmonella enteritidis*, *Acinetobacter baumannii*, *Citrobacter freundii*, *Enterobacter cloacae*, and *Klebsiella* species. Within Grampositive microorganisms, *Clostridium perfringens* is most commonly observed. Previous research has linked biliary infection with gallstone development and indicated that bacteria may act as the nucleating factor initiating the formation of both pigment and cholesterol gallstones. Many studies [22, 23] had established the coexistence of biofilm-forming bacteria in bile and gallbladder/gallstones (*Pseudomonas aeruginosa, E. coli, Klebsiella pneumoniae, Enterococcus* spp. and *Acinetobacter* spp.) in different combinations, and the presence of *Capnocytophaga* spp., *Lactococcus* spp., *Bacillus* spp., *Staphylococcus haemolyticus*, *Enterobacter* or *Citrobacter* spp., *Morganella* spp., *Salmonella* spp., and *Helicobacter pylori*.

All of the microbiological studies that led to the selection of these antibiotic regimens were carried out using standard culture methods. Recent studies of microbial detection by culture- vs. culture-free identification of microbial DNA by next-generation sequencing (NGS) for various purulent diseases have shown that traditional culture only identifies a portion of the bacteria present. Additionally, in some Asian countries, the presence of *H. pylori* has been detected infrequently in the gallbladder by PCR. Other molecular tools like RAPD fingerprinting, *cagA* gene detection, which represent a good marker for genome-sequencing projects, are available nowadays to detect the microbial strains causing infections in AC patients [24].

### **9.2 ESKAPE pathogens and role of bile in development of drug resistance**

Bile has bactericidal activity. However, many pathogens are known to resist the bactericidal activity of bile and utilize this host component as a localization signal to regulate virulence gene expression and enhance infection. Furthermore, strategies employed by pathogens to resist bile align with antibiotic resistance

mechanisms. The efflux pump genes, *acrAB* in *E. coli*, *Salmonella*, *Shigella*, *Klebsiella*, and other pathogens, resist both bile salts and antibiotics, thereby making it essential for survival under extreme environmental conditions [25–28].

The ESKAPE group of pathogens (*Enterococcus, Staphylococcus, Klebsiella, Acinetobacter, Pseudomonas, and* Enterobacter) represents a significant public health threat as antibiotic resistance rates rise from the acquisition of multiple resistance mechanisms involving the gene expression of BSH, Gls24, GlsB, EmrB/QacA, PrkC, LiaFSR, BsrXRS, MnhF, WTA, OxyR, CpxAR, KpnO, KpnEF, CadC, TdcA, Gal ET, *pgaABCD*, *pqsABCDE*, ExoU, T6SS genes or through biofilm production, etc. Many ESKAPE pathogens are not known to cause infection in the gastrointestinal system; nevertheless, isolation from bile, the gallbladder, pancreatic or biliary stent biofilms, and bile duct infections have been described, and antibiotic resistance is frequently identified. Given a previous research that found positive bile cultures in 22.2% of the cases following elective gallbladder removal surgery, the findings are not restricted to hospital-based infections. Enterococcus spp. was the most frequent bacterial isolate found in the bile samples, followed by *E. coli*, *Klebsiella* spp., *Enterobacter* spp., and *Pseudomonas* spp.; 22.7% of these isolates were antibioticresistant. Furthermore, bile exposure in the lungs of CF patients has been shown to affect *Staphylococcus aureus, A. baumannii,* and especially *P. aeruginosa* infection. Growth was either consistent or increased in the presence of human bile, respectively, for *E. coli* or *Enterococcus faecalis*. Furthermore, bile reduced the antimicrobial activity of ciprofloxacin, meropenem, and tigecycline for *E. coli*, while linezolid and tigecycline had reduced activity against *E. faecalis* [29].

#### **9.3 Rare cases of acute cholecystitis caused by microorganisms**

Berinson et al. [30] reported one rare case of AC caused by *Kosakonia cowanii*, formerly known as *Enterobacter cowanii*, which is a Gram-negative bacillus belonging to the order *Enterobacterales*. The species is usually recognized as a plant pathogen and has only anecdotally been encountered as a human pathogen. A cholecystectomy confirmed the diagnosis of acute cholecystitis with partial gall bladder necrosis. By MALDI-TOF, 16S-rRNA analysis, and whole-genome sequencing, a surgical material produced pure cultures of Gram-negative rods that were clearly identified as K. cowanii.

Deering et al. [31] reported a rare case of acute cholecystitis caused by *Streptococcus bovis* biotypes (I & II), a Gram-positive, catalase-negative, anaerobic coccus found as a commensal inhabitant of the digestive system in 16% of healthy people. The patient was treated with tazobactam/piperacillin and later on subjected to laparoscopic cholecystectomy.

Vogt et al. [32] reported isolated Serogroup O1 *Vibrio cholerae* in an 83-year-old man suffering from AC. The Gram stain of the body fluid specimen demonstrated rare Gram-negative rods and many polymorphonuclear lymphocytes. The organism was positive for oxidase, and the results obtained using a Neg Breakpoint Combo Panel Type 41 (NBC41) and a MicroScan WalkAway Plus system (Siemens Healthcare Diagnostics, Deerfield, IL) identified the organism as *V. cholerae*, with 97.76% probability. The isolate was also tested using a manual API 20E Gramnegative identification panel (bioMerieux, Inc.,), which yielded a code of 5,347,124, giving a presumptive identification of *V. cholerae* at 99.9%.

#### **9.4 Viral causes of biliary tract infections**

In comparison with bacterial infections, viral infections of the biliary tract are less common and less discussed. Viral infections frequently occur as a result of a

*Infections of Biliary Tract DOI: http://dx.doi.org/10.5772/intechopen.100063*

liver infection or as part of a systemic viral illness. Viruses seldom cause primary liver infection. Cholangitis, or inflammation of the bile duct, is a very frequent symptom. Despite the fact that hepatotropic viruses (A, B, C, and E) are commonly thought of as hepatocellular pathogens, cholangitic symptoms are now widely documented in conjunction with these disorders [10, 14, 23, 33]. Cholangitis is also due to systemic viral infections in different proportions to hepatitis. The human immunodeficiency virus (HIV) is linked to a variety of liver problems, including cholangitis. Other systemic viruses, most notably members of the herpes virus family, can induce hepatic illness in both immunocompromised and immunocompetent individuals, including cholangitis and potentially ductopenia [34].

#### **9.5 Parasitic causes of biliary tract infections**

Cholangitis can be caused due to a variety of reasons, including biliary calculi, strictures, parasites, post-endoscopic retrograde cholangiopancreatography (ERCP), postoperative, and so on. Biliary parasitoses, in contrast to other causes, are more prevalent in many nations. Ascaris lumbricoides, liver flukes, and Echinococcus are common parasites that affect the biliary system*.* The trematodes (flukes) that commonly infect the human biliary tract include *Clonorchis sinensis, Opisthorchis viverrini, Opisthorchis felineus, and Fasciola hepatica.* The majority of patients are asymptomatic. While entering through the bile duct, they cause biliary colic and obstructive jaundice. The parasites reside in the intrahepatic bile ducts and, occasionally, in the extrahepatic bile ducts, gallbladder, and pancreatic duct. The result is mechanical obstruction, inflammatory reaction, adenomatous hyperplasia, and periductal fibrosis. The parasite can be examined through radiological findings of CT and MRI [35].

#### **9.6 Diagnosis**

The diagnostic criteria include examining for signs of local inflammation, such as Murphy's sign, the presence of a mass, pain, or tenderness located in the upper right quadrant of the abdomen. The local inflammation is often accompanied by systemic inflammation, indicated by signs of fever, increased white blood cell (WBC) counts, and elevated levels of C-reactive protein. The severity of acute cholecystitis can range from mild and self-limiting to severe and potentially life threatening [36, 37]. Several imaging techniques such as ultrasonography, magnetic resonance imaging (MRI), computed tomography (CT) are necessary to accurately diagnose both the typical and atypical cases of acute cholecystitis. Recently, Amini et al. had used high mobility group box protein 1 (HMGB1) biomarker for acute cholecystitis diagnosis [38].

#### *9.6.1 Diagnosis of cholecystitis*

For the consensus in diagnosis of cholecystitis in 2007, the Tokyo guidelines for the management of acute cholangitis and cholecystitis (TG07) were formed and widely adopted. In 2013, the updated Tokyo guidelines (TG13) for acute cholangitis and acute cholecystitis were released for severity grading of acute cholecystitis [37] (**Table 1**).

#### *9.6.2 TG07 severity assessment criteria*

The severity assessment criteria were first presented throughout the world in TG07 by Hirota and Takada, [37] where the severity grading of acute cholecystitis


#### **Table 1.**

*TG13 diagnostic criteria for acute cholecystitis.*

was classified into the following three categories: "mild (Grade I)," "moderate (Grade II)," and "severe (Grade III)."

Mild (Grade I) acute cholecystitis occurred in a patient with no signs of organ failure and mild gallbladder illness, allowing cholecystectomy to be performed safely and with minimal risk. The severity score for these individuals in TG07 does not fulfill the criteria for "moderate (Grade II)" and "severe (Grade III)" acute cholecystitis.

Acute cholecystitis, in which the degree of acute inflammation is expected to be linked with greater operating difficulties in completing cholecystectomy, was classified as moderate (Grade II) acute cholecystitis [8, 9, 16].

Severe (Grade III) acute cholecystitis was defined as acute cholecystitis associated with organ dysfunction (**Table 2**).

Reference: Masamichi et al. [19].

#### **9.7 Treatment**

Acute cholecystitis is often treated promptly by cholecystectomy or percutaneous cholecystostomy and antibiotic therapy in high-risk patients. Antimicrobial treatment has a different role depending on the severity of the illness and its etiology. Because it is unclear if bacteria have a role in grade I acute cholecystitis, antimicrobial treatment is used to prevent infection before cholecystectomy. Antimicrobial treatment is therapeutic and necessary for grade II acute cholecystitis until the gallbladder is removed. Most patients with bacteremia might have clinical deterioration and can be classified as grade III acute cholecystitis and are therefore not suitable for surgery. A recent meta-analysis reported that cholecystography has the highest diagnostic accuracy for detection of acute cholecystitis [39].

Previous studies have found bile to be infected in 9–42% of patients who underwent elective laparoscopic cholecystectomy, but the incidence of culture-positive bile increased to 35–65% of patients with acute cholecystitis [40]. Antimicrobial treatment is critical for reducing both the systemic septic response and local inflammation following cholecystectomy in individuals with moderate-to-severe acute cholecystitis [41]. Those with septic shock should get appropriate antibiotic treatment within 1 hour of diagnosis, and patients who are less severely sick should receive it within 6 hours. Bile culture results, however, cannot be acquired promptly after admission, and bile culture necessitates percutaneous gallbladder puncture. As a result, the most successful empiric antibiotics described in the literature are used as the basis for first antimicrobial treatment [42].

#### **Associated with dysfunction of any one of the following organ/systems**


1. Elevated white blood cell count ([18,000/mm3 )

2. Palpable tender mass in the right upper abdominal quadrant

3. Duration of complaints (72 h)

4. Marked local inflammation (gangrenous cholecystitis, pericholecystic abscess, hepatic abscess, biliary peritonitis, and emphysematous cholecystitis)

#### **Grade I (mild) acute cholecystitis**

Does not meet the criteria of "Grade III" or "Grade II" acute cholecystitis. Grade I can also be defined as acute cholecystitis in a healthy patient with no organ dysfunction and mild inflammatory changes in the gallbladder, making cholecystectomy a safe and low-risk operative procedure.

#### **Table 2.**

*TG 13 severity grading for acute cholecystitis.*

Because most infections in acute cholecystitis are limited to the gallbladder, sampling should be done directly from the infection site in order to identify the true causative pathogen. Bile specimens collected from the biliary tract using percutaneous transhepatic biliary drainage (PTBD) or endoscopic nasobiliary drainage (ENBD) are potentially associated with microbial contamination [43].

Bacterial infection is commonly reported in 50 to 90% of the cases. Most of the studies reported the involvement of polymicrobial infections in AC, which were often treated with antibiotic regimens with two or more antibiotics, but only one study had reported that monomicrobial growth was involved in AC. The most common presumptive antibiotics used in AC are ceftriaxone (2gm, IV, OD) or piperacillin/tazobactam (4.5 gm, IV, 8 hourly) or cefoperazone/sulbactam (3gm, IV, 12 hourly) for 7 to 10 days. The second-line or alternative antibiotics is imipenem (500 mg, IV, 6 hourly) or meropenem (1gm, IV, 8hourly) for 7 to 10 days. The most commonly isolated microorganisms among pathogens in positive bile cultures are Enterococci species, non-*faecium* enterococci (*Enterococcus faecalis*, *Enterococcus gallinarum*, *Enterococcus casseliflavus*, *Enterococcus avium*), *Escherichia Coli*, and *Klebsiella* species [44]. Gram-positive microbes, such as Enterococci, have become less common over time, whereas Gram-negative germs, particularly Enterobacteriales, have become more common and are most typically isolated among patients with acute cholecystitis. The results of local antimicrobial susceptibility tests, as well as information of the likely infecting microorganisms, pharmacokinetics/pharmacodynamics, and adverse reactions/effects of available medicines, must all be considered when making antimicrobial therapy decisions (local antibiogram). The severity of the illness and previous antimicrobial exposure are also important considerations in deciding the best course of treatment. β-lactam antibiotics or their derivatives, cephalosporins, carbapenems, fluoroquinolones,

and other antibiotics diminish infection. For moderate and severe acute cholecystitis, empiric treatment with piperacillin/tazobactam or a cephalosporin with or without metronidazole is advised, regardless of whether or not there is growth on culture [45].

Broad-spectrum β-lactam and β-lactamase inhibitors, such as ampicillinsulbactam, have been recommended as the first-line drugs to treat Enterococci and non-*faecium* enterococci infections. However, these microorganisms are reported to be resistant to most of the classes of antibiotics represented earlier. VREFM (vancomycin-resistant *Enterococcus faecium*) was reported for the first time in 2021 by Suk-Won et al. [46]. The authors found that the majority of the patients were suffering from Grade II acute cholecystitis (94.7%). Hence, they recommended other antibiotics, such as linezolid and tigecycline, which provide good coverage against VREFM, should be considered for patients with such advanced infections. Tigecycline can be used in several other cases because of its broad spectrum of effectiveness against Gram-negative microorganisms, including ESBL-producing bacteria. Tasina et al. [47] reported poor effectiveness of tigecycline toward a severely ill patient with AC.

Piperacillin-tazobactam and third- or fourth-generation cephalosporins are indicated as first-line antibiotics for Gram-negative bacteria, with fluoroquinolones and carbapenems as second-line antibiotics, depending on the severity of the infection and antimicrobial susceptibility patterns. According to Gomi et al. [48], most identified strains were resistant to ciprofloxacin due to widespread use of the antibiotic by the community, whereas 20% of pathogenic bacteria were resistant to ceftriaxone. As a result, in such circumstances, piperacillin-tazobactam or cefepime, which have larger spectra and lower resistance rates, are indicated. Carbapenem and tigecycline are advised for patients who are taking antibiotics on a regular basis. However, because of widespread medication resistance and associated high morbidity and mortality rates, carbapenem-resistant strains (CRE) species have emerged as major healthcare-related diseases [49].

#### *9.7.1 Empiric antibiotic treatment of community-acquired biliary tract infections (CA-BTI)*

The most important approach in controlling the CA-BTI is the primary source controls such as biliary drainage, removal of biliary tract stones, and cholecystectomy. The primary source control can help the antibiotics to penetrate the biliary tract, resulting in a better bactericidal effect when biliary obstruction is present. While it comes to medical therapy, there are two crucial variables to consider when choosing empiric antibiotics. Administration of antibiotics is essential for the treatment of BTI, in addition to primary source control. As the BTI is caused by endogenous etiological agents, that is, gastrointestinal tract flora, such as Escherichia coli, Klebsiella spp., Enterococci spp., Bacteroides spp., antibiotics that are effective against these organisms are usually used empirically to treat BTI rather than definite therapy. However, the usage of inappropriate empiric antibiotics may also incur fatal outcomes. To elicit positive treatment responses, >80% of the presumed causative microorganisms should be sensitive to antibiotics, and for patients with septic shock, the susceptibility rates should even exceed 100% [50]. Next, the antibiotics must be present in adequate concentrations at the infection sites to have the desired antimicrobial action [51, 52]. **Table 3** shows the antibiotics usually used to treat biliary tract infections based on their biliary penetration ability (indicated by the ratio of bile-to-serum concentrations [53–55].

Augmentin = amoxicillin + clavulanate; Bile/serum = bile concentration/serum concentration; Tazocin: Piperacillin + tazobactum; Unasyn = ampicillin + sulbactum.


#### **Table 3.**

*Antibiotics frequently used to treat biliary tract infections and their biliary penetration ability (indicated as the ration of bile to serum concentrations).*

As a result, when choosing empiric antibiotics for the treatment of BTI, both susceptibility rates and the potential of biliary penetration should be taken into account. **Table 3** lists the antibiotics often used to treat BTI, as well as their biliary penetration ability (measured as the ratio of bile-to-serum concentrations). Only individuals with a reasonable ratio (>1) of bile-to-serum concentrations (**Table 3**) could be candidates for empiric antibiotics for BTI, according to the criteria outlined earlier. The local antimicrobial susceptibility patterns of the usual causative agents for BTI should also be considered when prescribing appropriate empiric antibiotics. To ensure a positive outcome, only those with a 20% resistance rate should be used as empirical antibiotics.

Patients with severe cholecystitis are unfortunately difficult to identify effectively, both clinically and radiologically, because clinical presentations are unpredictable, and imaging findings are frequently ambiguous. However, there are significant differences in morbidity and fatality rates between patients with uncomplicated cholecystitis and those with severe cholecystitis. Preventing related consequences requires early detection and careful management of patients at risk of severe cholecystitis.

#### **10. Conclusions**

When acute cholecystitis is suspected, bile samples are taken for microbiology culture and sensitivity testing, and antibiotics are prescribed once the diagnosis has been established. The antibiotics of choice are parenteral cephalosporin or ampicillin, as well as aminoglycosides. The antibiotic regimen chosen is based on the severity of the clinical presentation. Because acute suppurative cholangitis with biliary blockage has a high pre- and postoperative mortality rate, comprehensive antimicrobial therapy is required following biliary decompression. Bile microbiological analysis is an expedient diagnostic tool for determining more suitable medication and generating local antibiotic guidelines for the treatment of biliary tract infections.
