Agreement between microscopic examination and bacterial culture of bile samples for detection of bactibilia in dogs and cats with hepatobiliary disease

Medora B. Pashmakova Department of Small Animal Clinical Sciences, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, TX 77843.

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Julie Piccione Department of Veterinary Pathobiology, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, TX 77843.

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Micah A. Bishop Department of Small Animal Clinical Sciences, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, TX 77843.

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Whitney R. Nelson Department of Small Animal Clinical Sciences, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, TX 77843.

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Sara D. Lawhon Department of Veterinary Pathobiology, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, TX 77843.

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Abstract

OBJECTIVE To evaluate the agreement between results of microscopic examination and bacterial culture of bile samples from dogs and cats with hepatobiliary disease for detection of bactibilia.

DESIGN Cross-sectional study.

ANIMALS 31 dogs and 21 cats with hepatobiliary disease for which subsequent microscopic examination and bacterial culture of bile samples was performed from 2004 through 2014.

PROCEDURES Electronic medical records of included dogs and cats were reviewed to extract data regarding diagnosis, antimicrobials administered, and results of microscopic examination and bacterial culture of bile samples. Agreement between these 2 diagnostic tests was assessed by calculation of the Cohen κ value.

RESULTS 17 (33%) dogs and cats had bactibilia identified by microscopic examination of bile samples, and 11 (21%) had bactibilia identified via bacterial culture. Agreement between these 2 tests was substantial (percentage agreement [positive and negative results], 85%; κ = 0.62; 95% confidence interval, 0.38 to 0.89) and improved to almost perfect when calculated for only animals that received no antimicrobials within 24 hours prior to sample collection (percentage agreement, 94%; κ = 0.84; 95% confidence interval, 0.61 to 1.00).

CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that agreement between microscopic examination and bacterial culture of bile samples for detection of bactibilia is optimized when dogs and cats are not receiving antimicrobials at the time of sample collection. Concurrent bacterial culture and microscopic examination of bile samples are recommended for all cats and dogs evaluated for hepatobiliary disease.

Abstract

OBJECTIVE To evaluate the agreement between results of microscopic examination and bacterial culture of bile samples from dogs and cats with hepatobiliary disease for detection of bactibilia.

DESIGN Cross-sectional study.

ANIMALS 31 dogs and 21 cats with hepatobiliary disease for which subsequent microscopic examination and bacterial culture of bile samples was performed from 2004 through 2014.

PROCEDURES Electronic medical records of included dogs and cats were reviewed to extract data regarding diagnosis, antimicrobials administered, and results of microscopic examination and bacterial culture of bile samples. Agreement between these 2 diagnostic tests was assessed by calculation of the Cohen κ value.

RESULTS 17 (33%) dogs and cats had bactibilia identified by microscopic examination of bile samples, and 11 (21%) had bactibilia identified via bacterial culture. Agreement between these 2 tests was substantial (percentage agreement [positive and negative results], 85%; κ = 0.62; 95% confidence interval, 0.38 to 0.89) and improved to almost perfect when calculated for only animals that received no antimicrobials within 24 hours prior to sample collection (percentage agreement, 94%; κ = 0.84; 95% confidence interval, 0.61 to 1.00).

CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that agreement between microscopic examination and bacterial culture of bile samples for detection of bactibilia is optimized when dogs and cats are not receiving antimicrobials at the time of sample collection. Concurrent bacterial culture and microscopic examination of bile samples are recommended for all cats and dogs evaluated for hepatobiliary disease.

Microscopic examination and bacterial culture of bile samples are common during diagnostic work-up of hepatobiliary disease in small animals.1–4 The importance of detection of bacteria in the bile varies among species. Bile in healthy people and cats is reportedly sterile,5,6 whereas occasional presence of enteric bacteria in the bile of healthy dogs has been reported.7 In clinically ill animals, bactibilia may signify the presence of bacterial cholangitis and warrant therapeutic intervention.

The hepatobiliary system is equipped with several mechanisms that maintain bile sterility. Antegrade flow of bile and a functional sphincter of Oddi have mechanical functions, whereas the local presence of IgA and bacteriostatic bile salts have immunochemical roles.1 Additionally, tight junctions and Kupffer cells prevent bacterial translocation at the interface between bile and blood in the liver parenchyma. When these mechanisms are disrupted by disease, ascending or hematogenous spread of bacteria into the hepatobiliary tree can occur, resulting in considerable morbidity and death secondary to hepatobiliary sepsis.1

Identification of bactibilia in dogs and cats suspected to have bacterial cholangitis has the potential to affect clinical decisions regarding treatment, particularly when bacterial identification is confirmed by results of culture. Microscopic findings for bile samples can be available on the day of sample collection in facilities where a clinical pathologist is readily available, whereas bacterial culture of bile samples requires 3 to 5 days or longer. Furthermore, inappropriate storage and transport conditions of specimens for bacterial culture can compromise the results. For these reasons, understanding the relationship between detection of bactibilia via microscopy and detection via bacterial culture may be of clinical benefit.

The primary objective of the study reported here was to evaluate the agreement between results of microscopic examination and bacterial culture for detection of bactibilia by use of bile samples from dogs and cats with hepatobiliary disease that had undergone percutaneous cholecystocentesis for bile sample collection. The secondary objective was to evaluate the agreement between these 2 diagnostic tests in a subset of patients that received no antimicrobial treatment within the 24-hour period prior to bile sample collection.

Materials and Methods

Animals and data collection

Medical records were searched to retrospectively identify dogs and cats that had undergone ultrasound-guided percutaneous cholecystocentesis from 2004 through 2014 and for which concurrent microscopic examination and bacterial culture of bile samples had been performed. For each patient, data were extracted regarding signalment, diagnostic code words, antimicrobial administration 24 hours prior to bile sample collection, and results of microscopic examination (for bacteria and inflammatory cells) and bacterial culture (aerobic, anaerobic, or both) of bile samples, including the main 3 microorganisms isolated. Antimicrobial susceptibility testing data (reflecting susceptibility patterns at the authors’ institution) for all cats and dogs with positive results of bacterial culture were also retrieved.

Bile sample collection

For all included dogs and cats, ultrasound-guided cholecystocentesis had been performed from a transhepatic percutaneous approach5 with the patient sedated or anesthetized. A 22-gauge, 1.5-inch needle with an attached 12-mL syringe had been used. The syringe had been advanced until the tip was visible in the gallbladder lumen. A minimum yield of 1 to 3 mL of bile had been aspirated for diagnostic evaluation, and the sample had been submitted for microscopic examination and bacterial culture within 1 hour after collection.

Microscopic examination of bile samples

Direct and concentrated smears and cytocentrifuge preparations had been prepared from each collected bile sample. Prepared slides had been stained with modified Wright staina by use of an automated slide stainerb and examined by a board-certified clinical pathologist or resident supervised by a board-certified clinical pathologist for the presence of crystals, bacteria, inflammatory cells, and epithelial cells.

Results of these historical tests were classified for data acquisition and statistical purposes in the present study as consistent with normal bile without bacteria or inflammatory cells (ie, negative for bactibilia; Figure 1); presence of bacteria without inflammatory cells (ie, positive for bactibilia; Figure 2); presence of bacteria and inflammatory cells (ie, positive for bactibilia and positive for inflammatory cells); or septic bile (presence of inflammatory cells with phagocytized bacteria, with or without extracellular bacteria; positive for bactibilia and positive for inflammatory cells; Figure 3).8 Both categories involving the presence of both bacteria and inflammatory cells were considered as reflective of inflammatory bactibilia for statistical purposes.

Figure 1—
Figure 1—

Photomicrograph of a cytocentrifuge preparation of a cytologically unremarkable bile sample from a dog. Modified Wright stain; bar = 10 μm.

Citation: Journal of the American Veterinary Medical Association 250, 9; 10.2460/javma.250.9.1007

Figure 2—
Figure 2—

Photomicrograph of a cytocentrifuge preparation of a bile sample from a dog considered to have bactibilia. Several short chains of large cocci are visible in a background of bile. Modified Wright stain; bar = 10 μm.

Citation: Journal of the American Veterinary Medical Association 250, 9; 10.2460/javma.250.9.1007

Figure 3—
Figure 3—

Photomicrograph of a cytocentrifuge preparation of a bile sample from a dog considered to have septic bile. Numerous markedly degenerative neutrophils are visible, many of which contain bacteria. Modified Wright stain; bar = 10 μm.

Citation: Journal of the American Veterinary Medical Association 250, 9; 10.2460/javma.250.9.1007

Bacterial culture of bile samples

Bile samples for bacterial culture had been used to inoculate trypticase soy agar plates supplemented with 5% sheep blood (blood agar plate).c Bile samples had also been used to inoculate MacConkey agar platesc to select for gram-negative enteric bacteria and trypticase soy brothc for broth enrichment. The broth enrichment product had been subcultured onto a blood agar plate. For culture of aerobic organisms, inoculated plates had been incubated overnight (approx 18 hours) at 37°C in a 5% carbon dioxide atmosphere.d For culture of anaerobic organisms, bile samples had been used to inoculate Brucella agar plates supplemented with 5% sheep blood, hemin, and vitamin Ke as well as brain heart infusion brothc supplemented with an oxygen-reducing enzymef; plates and broth were incubated at 37°C in a reduced oxygen environment.g

Aerobic and anaerobic culture plates had been examined for bacterial growth frequency for up to 5 days. Antimicrobial susceptibilities of bacterial isolates had been determined with commercially available systemsh,i or by disk diffusion in accordance with the performance standards of the Clinical Laboratory Standards Institute.9

Statistical analysis

The Cohen κ test was used to measure agreement between microscopic examination and bacterial culture for detection of bactibilia by use of statistical software.j Agreement was considered slight when the κ value was < 0.2, fair when the value was 0.2 to 0.4, moderate when the value was 0.4 to 0.6, substantial when the value was 0.6 to 0.8, and almost perfect when the value was > 0.8.10 Percentage agreement between the 2 diagnostic tests in results (both positive and negative) was also calculated. Additional calculations were performed to determine the proportion of bile samples in which bacteria were identified microscopically that also contained inflammatory cells as well as the proportion of bile samples in which inflammatory bactibilia was identified microscopically that also yielded positive results of bacterial culture.

Results

Fifty-two animals (31 dogs and 21 cats) with various types of hepatobiliary disease met the study inclusion criteria (Table 1). Results of microscopic examination of bile samples revealed normal bile (without bacteria or inflammatory cells) for 35 animals, bacteria without inflammatory cells for 12 animals, bacteria and inflammatory cells for 3 animals, and septic bile for 2 animals. Seventeen (33%) animals were classified as having bactibilia.

Table 1—

Number (%) of 31 dogs and 21 cats with various diagnoses that underwent percutaneous cholecystocentesis with subsequent microscopic examination and bacterial culture of obtained bile samples.

DiagnosisDogsCats
Cholecystitis7 (23)1 (5)
Cholangitis3 (10)5 (24)
Cholangiohepatitis2 (6)4 (19)
Chronic active hepatitis3 (10)0 (0)
Undefined hepatopathy3 (10)4 (19)
Hepatic lipidosis0 (0)4 (19)
Enteropathy2 (6)0 (0)
Gallbladder mucocele1 (3)0 (0)
Liver failure1 (3)0 (0)
Copper hepatopathy1 (3)0 (0)
Toxic hepatopathy1 (3)0 (0)
Emphysematous cholecystitis1 (3)0 (0)
Cirrhosis, acquired portosystemic shunt1 (3)0 (0)
Cholelithiasis1 (3)0 (0)
Chronic liver disease with bridging portal fibrosis1 (3)0 (0)
Disseminated coccidioides1 (3)0 (0)
Unregulated diabetes with high liver enzyme activity1 (3)0 (0)
Vacuolar hepatopathy1 (3)0 (0)
Liver lymphoma0 (0)1 (5)
Ductal plate malformation (liver)0 (0)1 (5)
Biliary adenocarcinoma0 (0)1 (5)

For 8 animals, only aerobic culture of bile samples was performed; for the remaining 44 animals, both aerobic culture and anaerobic culture were performed. Eleven (21%) animals had positive results of bacterial culture (Table 2), and Enterococcus spp (n = 7) and Escherichia coli (6) were the most common isolates. Both Enterococcus spp and E coli were isolated from bile samples of 6 animals. Only 1 anaerobic organism (Parabacteroides distasonis) was recovered from a total of 44 anaerobic cultures. One bile sample for which microscopic examination revealed normal bile yielded growth of a Campylobacter sp. Microscopic examination of 5 bile samples revealed inflammatory cells, and 2 of these samples met the criteria for septic bile; all 5 samples were identified microscopically to contain bacteria. Only 1 of the 3 samples that were not classified as septic bile via microscopy yielded positive results of bacterial culture (2 E coli isolates and 1 Corynebacterium isolate). In some instances, multiple organisms were isolated from the same sample. Antimicrobial susceptibility testing results for selected isolates were summarized (Table 3).

Table 2—

Results of microscopic examination and bacterial culture of bile samples collected from 31 dogs and 21 cats with hepatobiliary disease.

Microscopic examination resultsNo. of samples with no bacterial growthNo. of samples with bacterial growthBacteria isolated (No. of samples)
Normal bile (n = 35)341Campylobacter sp (1)
Bacteria (n = 12)57
  • Escherichia coli (3)

  • Enterococcus spp (5)

  • Gram-positive pleomorphic rod (1)

  • α-Hemolytic Streptococcus sp (1)

  • Bacillus sp (1)

Bacteria and inflammatory cells (n = 3)21
  • E coli (2)

  • Corynebacterium sp (1)

Septic bile* (n = 2)02
  • E coli (1)

  • Enterococcus spp (2)

  • Methicillin-resistant Staphylococcus pseudintermedius (1)

  • Parabacteroides distasonis (1)

Bile sample in which inflammatory cells containing phagocytized bacteria were detected.

Microorganism was grown in anaerobic culture conditions.

Bacterial culture was performed for each bile sample; 8 samples underwent aerobic bacterial culture only, and the remaining 44 samples underwent both aerobic and anaerobic bacterial culture. In some instances, multiple organisms were isolated from the same sample; therefore, total number of isolates may not match total number of samples with bacterial growth.

Table 3—

Minimum inhibitory concentration of selected antimicrobials against Enterococcus and E coli isolates recovered from bile samples* of the dogs and cats in Table 2.

OrganismAmikacinAmoxicillin-clavulanic acidAmpicillinCefpodoximeChloramphenicolClindamycinEnrofloxacinTetracyclineTrimethoprim sulfamethoxazole
E coli≤ 4 (S)≤ 4 (S)4 (S)≤ 2 (S)> 16 (R)> 2 (R)≤ 0.5 (S)> 8 (R)≤ 0.5 (S)
E coli≤ 4 (S)8 (S)> 16 (R)≤ 2 (S)≤ 4 (S)> 2 (R)≤ 0.5 (S)≤ 1 (S)≤ 0.5 (S)
E coli≤ 4 (S)> 32 (R)> 16 (R)≤ 2 (S)8 (S)> 1 (R)≤ 0.5 (S)4 (S)≤ 0.5 (S)
E coli≤ 4 (S)> 32 (R)> 16 (R)> 16 (R)16 (I)> 1 (R)> 4 (R)4 (S)≤ 0.5 (S)
E coli≤ 4 (S)> 1 (R)> 1 (R)> 16 (R)≤ 4 (S)> 4 (R)≤ 0.25 (S)≤ 2 (S)≤ 0.5 (S)
E coli≤ 4 (S)> 1 (R)> 1 (R)≤ 2 (S)8 (S)> 4 (R)≤ 0.25 (S)4 (S)≤ 0.5 (S)
Enterococcus spp— (S)— (S)— (I)— (R)
Enterococcus spp— (S)— (I)— (R)— (S)
Enterococcus spp0.5 (S)8 (S)0.5 (S)≥ 16 (R)
Enterococcus spp0.25 (S)4 (S)≥ 2 (R)≤ 1 (S)
Enterococcus spp≤ 0.12 (S)2 (S)≥ 2 (R)≤ 1 (S)
Enterococcus spp≥ 16 (R)8 (S)1 (I)≤ 2 (S)
Enterococcus spp≤ 4 (S)1 (I)≥ 8 (R)

Letters in parentheses represent interpretations (S = susceptible, I = intermediate, and R = resistant).

Both Enterococcus spp and E coli were isolated from bile samples of 6 animals.

— = Antimicrobial susceptibility results not reported for enterococci against aminoglycosides, cephalosporins, sulfonamides, and clindamycin because of lack of clinical correlation, in accordance with the published guidelines of the Clinical and Laboratory Standards Institute.9

Tetracycline susceptibility is considered predictive of doxycycline susceptibility. The 2 Enterococcus isolates lacking minimum inhibitory concentration data but with susceptibility test results were tested by disk diffusion.

See Table 2 for remainder of key.

Agreement between the results of microscopic examination and bacterial culture of bile specimens for detection of bactibilia in all dogs and cats was substantial (percentage agreement, 85% (44/52); κ = 0.62; 95% confidence interval, 0.38 to 0.85). Agreement between these results when including only dogs and cats that received no antimicrobials within 24 hours prior to bile sample collection improved to almost perfect (percentage agreement, 95% [35/37]; κ = 0.84; 95% confidence interval, 0.61 to 1.00).

Of the 17 bile samples in which bacteria were identified microscopically, 5 also contained inflammatory cells (indicative of inflammatory bactibilia). Of the 5 bile samples in which inflammatory bactibilia was identified microscopically, 3 yielded positive results of bacterial culture.

Discussion

Results of the present study indicated substantial agreement between results of microscopic examination and bacterial culture of bile specimens from dogs and cats with hepatobiliary disease for detection of bactibilia. This agreement improved to almost perfect for dogs and cats that received no antimicrobials within the 24 hours prior to bile sample collection. As found in previous studies,2–4 the enteric bacteria Enterococcus spp and E coli were the most common bacterial species isolated from bile samples. These bacteria were frequently present in combination with other bacteria. Also as previously reported,4 infectious organisms were more commonly detected via microscopy than via bacterial culture, and inflammatory cells were lacking in most samples in which bacteria were identified microscopically.

Bacterial recovery rates achieved in the present study might have been improved by the modification of the culture methods used, particularly the inclusion of chocolate agar for culture of fastidious aerobic bacteria, use of thioglycolate broth supplemented with vitamin K and hemin for aerobic culture enrichment, and use of chopped-meat carbohydrate broth for culture of fastidious anaerobic bacteria. Use of an oxygen-reducing enzyme rather than anaerobic broth inoculation and incubation in an anaerobic chamber might have also resulted in a lower recovery rate for anaerobic bacteria.

The results reported here represented 11 years of bacterial culture results, and the standard operating procedures of the laboratory changed considerably during that period. During part of this period, an anaerobic chamber was used for incubation of anaerobic culture plates (2006 through 2009), but culture plates were not inoculated under anaerobic conditions. Of the 11 bile samples that yielded positive culture results, 5 were submitted for bacterial culture during that period and 4 samples were processed for both aerobic and anaerobic culture. No anaerobic bacteria were recovered from these 4 samples. Of the remaining 6 bile samples for which bacterial culture was performed after 2009 when jars were used, an anaerobe, P distasonis, was isolated from 1 sample. Although use of an anaerobic chamber for bile sample setup and incubation of culture plates is cumbersome and expensive, it may improve recovery rates of obligate anaerobes. Additionally, incubation of anaerobic culture plates for a minimum of 7 days may improve recovery of obligate anaerobes from bile samples.11

In patients in the present study in which bactibilia was detected microscopically on the day of bile sample collection, bacterial culture was likely to yield bacterial growth for which the antimicrobial susceptibility pattern could be determined. Inversely, bile samples in which no bacteria were detected microscopically were unlikely to yield positive results of bacterial culture. However, when low numbers of culturable bacteria exist in bile samples, microscopic examination may yield negative results because a minimum of 1,000 to 100,000 bacteria/mL of fluid is believed to be needed for detection on a prepared slide.12 Therefore, it is still recommended that bile specimens be submitted for both microscopic examination and bacterial culture to optimize the possibility of identifying bacterial pathogens in bile.

Several factors may explain a lack of microbial growth in bile samples in which bactibilia is identified microscopically. Microscopic detection of bacteria provides no information regarding bacterial viability. This is particularly relevant in patients treated with antimicrobials, in which nonviable organisms may be microscopically visible. Timing of bile sample collection in relation to antimicrobial administration or disease onset, route of antimicrobial administration, and use of time-dependent versus dose-dependent antimicrobials may also play a factor.13 These factors were not specifically accounted for in the data analysis of the present study but may be more clearly elucidated in a prospectively conducted study. False-negative results of bacterial culture are also possible when samples are collected or stored improperly. This is particularly true with anaerobic cultures.13 Bacterial cultures included in the present study had been initiated on the same day as sample acquisition, making storage errors unlikely.

One bile sample in which no bacteria were identified microscopically in the present study yielded growth of a Campylobacter sp, a known enteric microbe, on bacterial culture. Sample contamination was considered unlikely in this situation because standard sterile technique had been used, and this bacterium could have ascended from the gastrointestinal tract into the biliary tree. Therefore, a low bacterial load was considered a more likely explanation for the negative results of microscopic examination in this situation.

From a bile sample from a different patient (a 2-year-old neutered male cat with cholangiohepatitis), Staphylococcus pseudintermedius was isolated, which is not a typical inhabitant of the gastrointestinal tract. This sample also yielded growth of Enterococcus spp. Additional specimens of liver tissue and abdominal fluid had been collected from this patient, yielding growth of Enterococcus spp and methicillin-resistant S pseudintermedius. This patient had a complex medical history that included prior oronasal fistula repair, chronic vomiting with placement of an esophageal feeding tube, and recurrent pneumonia. Therefore, hematogenous spread of S pseudintermedius into the bile, liver, and abdominal fluid may have occurred.

Although microscopic examination and bacterial culture of bile specimens are common diagnostic tests, their relative sensitivities for bacterial detection in bile samples remain unknown. Additional diagnostic tests of rapid clinical usefulness may include Gram staining to determine the Gram reaction and morphology of bacteria, particularly when antimicrobial choices need to be made before results of culture and antimicrobial susceptibility testing become available. Advanced microbial identification techniques such as PCR assay may also be useful for detection and speciation of bacteria in bile samples, although PCR assay of blood samples was no more sensitive than bacterial culture in a study14 of dogs with bacteremia.

Although not a concern for the dogs and cats in the present study, the potential for contamination of a sample with skin flora should be considered when bacterial culture of the sample yields positive results but microscopic examination reveals no bacteria. Knowledge of gastrointestinal flora that may ascend into the biliary tree is important in the context of interpreting culture results, given that this is the most commonly recognized route of entry in dog and cats.1 The bacteria isolated in the present study were all known to be of enteric origin, with the exception of methicillin-resistant S pseudintermedius, and therefore were not necessarily suggestive of bile sample contamination.

In the study reported here, 12 bile samples in which bacteria were identified microscopically had no microscopic evidence of inflammation; however, all 5 samples in which inflammatory cells were identified also contained extracellular bacteria. A similar disparity between microscopic detection of bacteria and inflammatory cells in bile samples was identified in a larger-scale study.4 Also of interest was the low proportion of bile samples positive for inflammatory bactibilia that yielded positive results of bacterial culture. This finding may be explained as false-negative culture results, given that we can offer no plausible reason that inflammatory cells would interfere with bacterial growth. However, the chemical composition of bile can affect the presence of inflammatory cells and their function locally, given that bile has immunomodulatory functions, including suppression of leukocyte activation and phagocytosis.15 Although this possibility has not been extensively described in the veterinary literature, a case report16 exists of noninflammatory bactibilia in a dog with clinically important bacterial cholecystitis.16 In humans with bacterial cholecystitis, inflammatory cells may also be lacking in bile samples. However, the pathophysiologic nature of cholecystitis in humans is different from that in small animals and is typically associated with biliary obstruction secondary to cholelithiasis, precluding the ability to make direct comparisons between species.17,18

Virulence factors associated with certain bacteria may also influence whether inflammation develops in hosts. Many bacteria, such as Enterococcus faecalis, possess mechanisms to evade the host immune response and decrease the degree of neutrophil recruitment, such as capsular polysaccharides production and an extracellular gelatinase matrix, respectively. Research has also demonstrated that, through rapid mutation, E coli can develop adaptive mechanisms to evade macrophage engulfment in as little as 4 days.19

In both human and veterinary medicine, debate exists on whether the presence of microbes in the bile mandates treatment.6 Additionally, Enterococcus spp are not always considered pathogenic. Although an in-depth discussion on antimicrobial treatment of biliary infections is beyond the scope of the present report, several guidelines may be prudent for clinical consideration. In general, the decision to treat a biliary infection should be made in the context of the likelihood that hepatobiliary disease is the cause of the observed clinical signs, illness severity, patient immunocompetency, and whether another identifiable source of infection or sepsis exists. Judicious empirical use of antimicrobials may be indicated in dogs and cats with bactibilia and signs of systemic infection. In those circumstances, treatment should be initiated immediately while awaiting definitive culture results and escalated or de-escalated appropriately as soon as culture results are available. Initial antimicrobial selection should be supported by the microscopic detection of pathogens in bile samples, bacterial characteristics identified via gram-stain analysis, and regional data regarding antimicrobial susceptibility patterns for the most likely pathogens (Enterococcus spp and E coli) until culture results become available. Conversely, antimicrobial treatment may be unnecessary in patients in which bactibilia is an incidental finding or is unaccompanied by signs of systemic disease.

The present study had several limitations. The low number of dogs and cats precluded analysis of species differences in bacterial isolate patterns or diagnostic test agreement, which would be interesting to discern in future studies. Diagnostic code words, timing of bile sample collection, and treatments were not standardized in a way that would allow analysis of their influence on the results. Likewise, ultrasonographic appearance of the hepatobiliary system may have offered additional insight, given that detection of biliary sludge is reportedly 100% specific for the diagnosis of bactibilia.2 However, over the study period, radiologic reports differed considerably with respect to terminology used in describing gallbladder appearance. Lastly, reliance on medical records rather than prospective recruitment of patients resulted in the inability to assess other variables such as patient history or therapeutic interventions on the study results.

Regardless of the aforementioned limitations, findings of the present study indicated substantial agreement between microscopic examination and bacterial culture of bile specimens for detection of bactibilia in dogs and cats with hepatobiliary disease. Enterococcus spp and E coli were the most common bacterial isolates. Because of the possibility that low bacterial counts may result in a lack of microscopic detection of bacteria in bile samples, we recommend that bacterial culture of bile samples be performed concurrently. Both diagnostic and clinical data should be judiciously interpreted to help guide clinical decisions regarding empirical antimicrobial treatment of individual dogs and cats.

Acknowledgments

Presented in abstract form at the American College of Veterinary Internal Medicine Forum, Indianapolis, June 2015.

The authors thank Kay Duncan for assistance with data retrieval for the bacterial cultures.

Footnotes

a.

ELITechGroup Inc, Logan, Utah.

b.

Aerospray Pro, ELITechGroup Inc, Logan, Utah.

c.

Becton, Dickinson & Co, Franklin Lakes, NJ.

d.

Thermo Scientific Corp, Marietta, Ohio.

e.

Remel, Thermo Scientific Corp, Marietta, Ohio.

f.

Oxyrase, Oxyrase Inc, Mansfield, Ohio.

g.

Mitsubishi AnaeroPack, Mitsubishi Gas Chemical, Tokyo, Japan.

h.

TREK Sensititre, TREK Diagnostics, Cleveland, Ohio.

i.

Vitek, bioMérieux, Durham, NC.

j.

QuickCalcs quantify agreement with kappa, GraphPad Software, La Jolla, Calif. Available at: www.graphpad.com/quickcalcs/kappa1.cfin. Accessed Nov 3, 2014.

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