Liver abscesses (LA) in feedlot cattle are polymicrobial infections, initiated primarily by the translocations of the two subspecies of Fusobacterium necrophorum, necrophorum and funduliforme, and secondarily by Trueperella pyogenes and Salmonella enterica from the rumen into the liver via portal blood. The hindgut has not been investigated as a potential source of LA pathogens. Our objectives were to analyze LA and matched ruminal and colonic epithelial tissues ( n = 96), collected at a commercial beef processor immediately after evisceration, to determine culture-based prevalence of pathogens, specifically, Escherichia coli , Klebsiella pneumoniae , Pseudomonas aeruginosa , and Bacteroides fragilis, which have not been targeted before, in addition to F. necrophorum, T. pyogenes, and S. enterica, and quantitative PCR-based prevalence and concentrations of the F. necrophorum subspecies. The subsp. necrophorum was the most dominant species in LA detected by both methods, and concentrations were in the range of 7.0 log ₁₀ colony forming units (CFU). Fusobacterial concentrations were higher in ruminal epithelial than in colonic epithelial tissues, but colonic epithelial tissues yielded more subsp. necrophorum isolates than ruminal epithelial tissues. Of the targeted bacterial species, E. coli was the most dominant, and none of the tissues was positive for B. fragilis . A majority of the isolations of E. coli were after enrichment, suggesting the concentrations were too low to contribute to the abscess development. The data reaffirm that LA are polymicrobial infections, F. necrophorum, specifically necrophorum, is the most dominant species prevalent, and the hindgut is likely to be a source, in addition to the rumen, of pathogens that cause LA. IMPORTANCE: Liver abscesses (LA) in beef cattle are initiated by entry of bacterial pathogens from the rumen via portal blood. The two most frequently isolated pathogens are the two subspecies of Fusobacterium necrophorum, followed by Trueperella pyogenes and Salmonella enterica . Our objectives were to determine the prevalence of pathogens that have been reported sporadically and have not been targeted before. Liver abscesses and matched ruminal epithelium and colonic epithelial tissues, collected from feedlot cattle at slaughter, were analyzed by culture and PCR methods. The subsp. necrophorum was the most dominant bacterium in liver abscesses. Colonic epithelial tissues yielded more subsp. necrophorum than ruminal epithelial tissues. Although Escherichia coli was the second most prevalent species, a majority of the isolates were obtained after enrichment, indicating low concentrations. Our data reaffirm that F. necrophorum is the dominant species prevalent in LA, and the hindgut is likely to be another source of pathogens that cause LA.

Liver abscesses (LA) in cattle are a polymicrobial infection, and the major bacterial pathogens associated are as follows: Fusobacterium necrophorum subsp. necrophorum (FNN), F. necrophorum subsp. funduliforme (FNF), Trueperella pyogenes (TP), and Salmonella enterica (SE). In polymicrobial infections, the contributions of individual species are difficult to assess. We hypothesized that species abundance in abscesses may be indicative of their contributions. Therefore, the objective was to develop a 4-plex quantitative PCR (qPCR) assay to determine prevalence and concentrations of the pathogens in LA ( n = 384) and matched ruminal (RT; n = 374) and colonic epithelial (CT; n = 256) tissues. Fusobacterium necrophorum , either FNN or FNF, was detected in 85.9% of LA by qPCR, which was slightly lower than the culture-based prevalence (89.1%). Only 16.9% of LA were positive for FNF with no FNN. The concentrations of FNN or FNF were 7 to 7.5 log ₁₀ CFU/g. The qPCR assay identified more ( P < 0.01) samples (29.2%) as positive for TP than the culture method (16.7%). The mean concentration of TP was 5.9 log ₁₀ CFU/g. None of the LA was positive for SE by the qPCR assay. The prevalence of FN was greater ( P < 0.01) in RT than CT (73.2% vs. 16%). The concentrations were in the 4 to 5 log ₁₀ CFU/g. The low concentrations of TP suggest that it is unlikely to be the primary etiologic agent. Although SE was detected by culture method, it was not quantifiable, which suggests that the species does not contribute to the development of LA. IMPORTANCE: Liver abscesses (LA) in cattle are a polymicrobial infection, and four bacterial pathogens are implicated: Fusobacterium necrophorum subsp. necrophorum (FNN) and subsp. funduliforme (FNF), Trueperella pyogenes (TP), and Salmonella enterica (SE). In mixed infections, the species abundance may be indicative of their contributions. Our objective was to develop a quantitative PCR assay to determine prevalence and concentrations of the pathogens in LA ( n = 384) and matched ruminal (RT; n = 374) and colonic tissues (CT; n = 256). The dominant species in LA was FNN with a mean concentration of 7.0 log ₁₀ CFU/g. The subsp. FNF without FNN was prevalent in a small number of LA, with a mean concentration of 7.0 log ₁₀ CFU. The concentration of TP was 5.0 log₁₀ CFU, which suggests that it is unlikely to be the primary etiologic agent. Although S. enterica was prevalent in LA, none was quantifiable, which suggests that it is not contributing to LA.

This experiment aimed to determine the impact of growth-inhibitory compounds on F. necrophorum subsp. necrophorum (F.n.n.), a primary etiologic agent of liver abscesses in feedlot cattle, in an in vitro rumen fermentation system. The assessment of inhibition was by quantitative PCR. We hypothesized that the addition of growth inhibitory compounds would reduce F.n.n. concentrations. A 2 x 2 x 6 factorial arrangement of treatments was used. Factors included: 1) sodium lactate (0.0 or 100 mM); 2) bacteria (0 or 1x10⁸ CFU/ml of F.n.n.); and 3) 1 mL of test compound [Control (solvent with no additive); Tylosin: 5 μg/mL; Compound A; Compound B; Compound C; and Compound D]. Prior to morning feeding, rumen contents were collected from a single ruminally canulated steer fed a grain-based finishing diet. Rumen fluid was strained through 4 layers of cheesecloth, mixed with McDougall’s buffer (1:1 ratio; RFB), and 30 mL of the RFB was added into 100 mL-serum bottles containing the appropriate treatments. Serum bottles (n = 3 replicates per treatment) were flushed with CO₂ closed with rubber stoppers, crimped with aluminum seals, and incubated at 39ºC for 0, 6, 12, and 24 h. Following each incubation time point, a 2-mL aliquot was removed from each serum bottle under a stream of CO₂ to maintain anaerobic conditions. Immediately after sample collection, pH was determined, and DNA was extracted by boiling the sample, centrifugation, and purifying with a GeneClean Turbo DNA extraction kit. The concentration of F.n.n was determined by a quantitative PCR assay targeting the promoter region of the leukotoxin genes. Concentrations of F.n.n. from the RFB were used to adjust F.n.n. concentrations for all treatments at all time points. There was a lactate x test compound x incubation time interaction (P < 0.05) for F.n.n. concentration. In the absence of lactate, serum bottles containing tylosin or test compound treatments had decreased (P < 0.05) F.n.n. concentrations by 24 h post incubation. However, when sodium lactate was added, no treatments had F.n.n. concentrations less than the control treatment at 24 h post incubation (P > 0.05). These data suggest that compounds used in this experiment reduced the concentrations of F.n.n by 24 h post incubation in the absence of additional lactate. However, when lactate, a substrate for F.n.n, was added to the in vitro system, the reduction in F.n.n. growth was not observed.