Is bacterial DNA a surrogate marker of bacterial translocation in cirrhosis?
Authors: Tetsuji Fujita
Journal: Journal of Hepatology (2008)

I read with interest the excellent clinical study by Gonzalez-Navajas and colleagues [[1]Gonzalez-Navajas J.M. Bellot P. Frances R. Zapater P. Munoz C. Garcia P. et al.Presence of bacterial-DNA in cirrhosis identifies a subgroup of patients with marked inflammatory response not related to endotoxin.J Hepatol. 2008; 48: 61-67Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar]. Following their previous study showing that about 40% of patients with advanced cirrhosis had bacterial DNA in blood and ascites and indicating that repeated episodes of bacterial translocation (BT) evidenced by the detection of same bacterial DNA in consecutive blood samples are highly suspected in these patients [[2]Frances R. Benlloch S. Zapater P. Gpnzalez J.M. Lozano B. Munoz C. et al.A sequential study of serum bacterial DNA in patients with advanced cirrhosis and ascites.Hepatology. 2004; 39: 484-491Crossref PubMed Scopus (124) Google Scholar]. In this study the authors investigated whether the presence of bacterial DNA is associated with pro-inflammatory cytokine and nitric oxide responses independently of the endotoxin pathway. Forty-seven (33%) of 141 patients with cirrhosis and non-infected ascites showed the simultaneous presence of bacterial DNA in blood and ascites. Among these 47 patients, the first eight with bacterial DNA from Gram-positive cocci, the first eight with DNA from Gram-negative bacteria, and the first 16 patients without bacterial DNA were included in the study. Serum endotoxin and lipopolysaccharide-binding protein levels (LBP) in patients with Gram-positive bacterial DNA were similar to those in patients without bacterial DNA. Regardless of lower LBP and endotoxin levels in patients with Gram-positive bacterial DNA compared to those in patients with Gram-negative bacterial DNA, serum levels of tumour necrosis factor (TNF)-α and interleukin-6 (IL-6) in patients with Gram-positive bacterial DNA were significantly higher than those in patients without bacterial DNA and equivalent to patients with Gram-negative bacterial DNA. The authors concluded that the detection of bacterial DNA allows the identification of BT in patients with advanced cirrhosis, which is associated with increased inflammatory responses independently of endotoxin signaling.The results of this study are consistent with the suggestion by a recent review [[3]Wiest R. Garcia-Tsao G. Bacterial translocation (BT) in cirrhosis.Hepatology. 2005; 41: 422-433Crossref PubMed Scopus (550) Google Scholar] that Gram-positive bacteria or their components play an important role in the inflammatory response in patients with cirrhosis because the expression of Toll-like receptor (TLR)-2, which responds predominantly to cell wall components of Gram-positive bacteria, has repeatedly been shown to be upregulated, whereas the expression of TLR-4, a specific transducing receptor for endotoxin, is unaltered or downregulated in cirrhosis. I would like to address three questions. First, I would like to ask the authors whether the relationship between the detection of bacteria by blood culture and the identification of bacterial DNA by polymerase chain reaction has previously been examined in cirrhotic or other patients. It has been reported that there were 16.6% false-positive results when DNA was isolated from healthy volunteers with a commercially available kit [[4]Heininger A. Binder M. Ellinger A. Pfisterer J. Botzenhart K. Unertl K. et al.Effect of comprehensive validation of the template isolation procedure on the reliability of bacteraemia detection by a 16S rRNA gene PCR.Clin Microbiol Infect. 2004; 10: 452-458Abstract Full Text Full Text PDF PubMed Scopus (10) Google Scholar]. Contamination can originate from mastermix reagents and the DNA extraction procedure. Second, the values of TNF-α and IL-6 reported in this study (mean TNF-α: 324.93 μg/mL, mean IL-6: 294.96 μg/mL in patients with bacterial DNA) are more than 1,000,000-fold higher than those of our data and the results of a recent study (mean TNF-α: 90.9 pg/mL, mean IL-6: 84.8 pg/mL in patients with decompensated cirrhosis) [[5]Elsing C. Harenberg S. Stremmel W. Herrmann T. Serum levels of soluble Fas, nitric oxide and cytokines in acute decompensated cirrhotic patients.World J Gastroenterol. 2007; 13: 421-425PubMed Google Scholar]. I think there is an error in the conversion of the units. Third, I would like to ask the authors whether the difference (<0.3 EU/mL) in endotoxin concentrations between the patients with Gram-negative bacterial DNA and those with Gram-positive bacterial DNA is clinically relevant. The biological activity of endotoxin is not only dependent on endotoxin and LBP concentrations but also influenced by soluble CD14 levels, lipoprotein concentrations, the expression of membrane CD14, TLR-4, MD-2, and the activity of transcription factors. I read with interest the excellent clinical study by Gonzalez-Navajas and colleagues [[1]Gonzalez-Navajas J.M. Bellot P. Frances R. Zapater P. Munoz C. Garcia P. et al.Presence of bacterial-DNA in cirrhosis identifies a subgroup of patients with marked inflammatory response not related to endotoxin.J Hepatol. 2008; 48: 61-67Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar]. Following their previous study showing that about 40% of patients with advanced cirrhosis had bacterial DNA in blood and ascites and indicating that repeated episodes of bacterial translocation (BT) evidenced by the detection of same bacterial DNA in consecutive blood samples are highly suspected in these patients [[2]Frances R. Benlloch S. Zapater P. Gpnzalez J.M. Lozano B. Munoz C. et al.A sequential study of serum bacterial DNA in patients with advanced cirrhosis and ascites.Hepatology. 2004; 39: 484-491Crossref PubMed Scopus (124) Google Scholar]. In this study the authors investigated whether the presence of bacterial DNA is associated with pro-inflammatory cytokine and nitric oxide responses independently of the endotoxin pathway. Forty-seven (33%) of 141 patients with cirrhosis and non-infected ascites showed the simultaneous presence of bacterial DNA in blood and ascites. Among these 47 patients, the first eight with bacterial DNA from Gram-positive cocci, the first eight with DNA from Gram-negative bacteria, and the first 16 patients without bacterial DNA were included in the study. Serum endotoxin and lipopolysaccharide-binding protein levels (LBP) in patients with Gram-positive bacterial DNA were similar to those in patients without bacterial DNA. Regardless of lower LBP and endotoxin levels in patients with Gram-positive bacterial DNA compared to those in patients with Gram-negative bacterial DNA, serum levels of tumour necrosis factor (TNF)-α and interleukin-6 (IL-6) in patients with Gram-positive bacterial DNA were significantly higher than those in patients without bacterial DNA and equivalent to patients with Gram-negative bacterial DNA. The authors concluded that the detection of bacterial DNA allows the identification of BT in patients with advanced cirrhosis, which is associated with increased inflammatory responses independently of endotoxin signaling. The results of this study are consistent with the suggestion by a recent review [[3]Wiest R. Garcia-Tsao G. Bacterial translocation (BT) in cirrhosis.Hepatology. 2005; 41: 422-433Crossref PubMed Scopus (550) Google Scholar] that Gram-positive bacteria or their components play an important role in the inflammatory response in patients with cirrhosis because the expression of Toll-like receptor (TLR)-2, which responds predominantly to cell wall components of Gram-positive bacteria, has repeatedly been shown to be upregulated, whereas the expression of TLR-4, a specific transducing receptor for endotoxin, is unaltered or downregulated in cirrhosis. I would like to address three questions. First, I would like to ask the authors whether the relationship between the detection of bacteria by blood culture and the identification of bacterial DNA by polymerase chain reaction has previously been examined in cirrhotic or other patients. It has been reported that there were 16.6% false-positive results when DNA was isolated from healthy volunteers with a commercially available kit [[4]Heininger A. Binder M. Ellinger A. Pfisterer J. Botzenhart K. Unertl K. et al.Effect of comprehensive validation of the template isolation procedure on the reliability of bacteraemia detection by a 16S rRNA gene PCR.Clin Microbiol Infect. 2004; 10: 452-458Abstract Full Text Full Text PDF PubMed Scopus (10) Google Scholar]. Contamination can originate from mastermix reagents and the DNA extraction procedure. Second, the values of TNF-α and IL-6 reported in this study (mean TNF-α: 324.93 μg/mL, mean IL-6: 294.96 μg/mL in patients with bacterial DNA) are more than 1,000,000-fold higher than those of our data and the results of a recent study (mean TNF-α: 90.9 pg/mL, mean IL-6: 84.8 pg/mL in patients with decompensated cirrhosis) [[5]Elsing C. Harenberg S. Stremmel W. Herrmann T. Serum levels of soluble Fas, nitric oxide and cytokines in acute decompensated cirrhotic patients.World J Gastroenterol. 2007; 13: 421-425PubMed Google Scholar]. I think there is an error in the conversion of the units. Third, I would like to ask the authors whether the difference (<0.3 EU/mL) in endotoxin concentrations between the patients with Gram-negative bacterial DNA and those with Gram-positive bacterial DNA is clinically relevant. The biological activity of endotoxin is not only dependent on endotoxin and LBP concentrations but also influenced by soluble CD14 levels, lipoprotein concentrations, the expression of membrane CD14, TLR-4, MD-2, and the activity of transcription factors.