In vitro antibacterial activity of doripenem against clinical isolates from French teaching hospitals: proposition of zone diameter breakpoints.

The aims of the study were to determine the in vitro activity of doripenem, a new carbapenem, against a large number of bacterial pathogens and to propose zone diameter breakpoints for clinical categorization in France according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) minimum inhibitory concentration (MIC) breakpoints. The MICs of doripenem were determined by the broth microdilution method against 1,547 clinical isolates from eight French hospitals. The disk diffusion test was performed (10-µg discs) according to the Comité de l'Antibiogramme de la Société Française de Microbiologie (CASFM) method. The MIC(50/90) (mg/L) values were as follows: methicillin-susceptible Staphylococcus aureus (MSSA) (0.03/0.25), methicillin-resistant Staphylococcus aureus (MRSA) (1/2), methicillin-susceptible coagulase-negative staphylococci (MSCoNS) (0.03/0.12), methicillin-resistant coagulase-negative staphylococci (MRCoNS) (2/8), Streptococcus pneumoniae (0.016/0.25), viridans group streptococci (0.016/2), ß-hemolytic streptococci (≤0.008/≤0.008), Enterococcus faecalis (2/4), Enterococcus faecium (128/>128), Enterobacteriaceae (0.06/0.25), Pseudomonas aeruginosa (0.5/8), Acinetobacter baumannii (0.25/2), Haemophilus influenzae (0.12/0.25), and Moraxella catarrhalis (0.03/0.06). According to the regression curve, the zone diameter breakpoints were 24 and 19 mm for MICs of 1 and 4 mg/L, respectively. This study confirms the potent in vitro activity of doripenem against Pseudomonas aeruginosa, Acinetobacter, Enterobacteriaceae, MSSA, MSCoNS, and respiratory pathogens. According to the EUCAST MIC breakpoints (mg/L) ≤1/>4 for Enterobacteriaceae, Pseudomonas aeruginosa, and Acinetobacter, and ≤1/>1 for streptococci, pneumococci, and Haemophilus, the zone diameter breakpoints could be (mm) ≥24/<19 and ≥24/<24, respectively.

[Activity of doripenem against anaerobic bacteria]. / Activité du doripénème sur les bactéries anaérobies strictes.

AIMS OF THE STUDY: This study examines the activity of doripenem, a new carbapenem compound compared with amoxicillin-clavulanic acid, piperacillin+tazobactam, imipenem, clindamycin and metronidazole against 316 anaerobes. METHODS: Inoculum preparation and agar dilution method were performed according to the CLSI method for anaerobes (M11A7). RESULTS: At a concentration of 4µg/ml doripenem and imipenem (IMP) inhibited 122 (96 %) and 126 (99 %) strains of the Bacteroides fragilis group, respectively. In contrast, doripenem appeared more potent than IMP against Gram-positive anaerobes inhibiting at the same concentration of 4µg/ml 145/145 strains (100 %) versus 115/145 for IMP (79.3 %). Against 316 anaerobic strains, the carbapenem doripenem had an MIC(50) of 0.25µg/ml and an MIC(90) of 2µg/ml. Results were similar to those for imipenem (MIC(50) of 0.125µg/ml and MIC(90) of 4µg/ml). If we consider the resistant breakpoints of the two carbapenems as defined by EUCAST, the resistance rate for doripenem (MIC>4µg/ml) 1.6 % is similar to that of imipenem (MIC>8µg/ml) 1.3 %. CONCLUSION: Thus independently of the PK/PD parameters the two carbapenems demonstrated very close activity; doripenem was more potent on Gram-positive anaerobes and slightly less potent against Gram-negative anaerobes mainly the B. fragilis group. Further clinical studies are needed to assess its usefulness in patients.

[Recommendations of the Infectious Disease Committee of the French Association of Urology (AFU): antibiotic prophylaxis for urological procedures]. / Recommandations de bonnes pratiques cliniques: l'antibioprophylaxie en chirurgie urologique, par le Comité d'infectiologie de l'association française d'urologie (CIAFU).

The French Association of anesthesiology (SFAR) has published in 1999 the Antibiotic prophylaxis guidelines. Antibiotic resistance has increased and new procedures appeared so new recommendations were needed. We present the antibiotic prophylaxis guidelines from the committee of infectious diseases of the French Association of Urology.

[Diagnosis and treatment of acute uncomplicated urethritis of men: guidelines from the French committee of infectious disease of the French association of urology (CIAFU)]. / Recommandations de bonnes pratiques cliniques : diagnostic et traitement des uréthrites aiguës non compliquées de l'homme, par le comité d'infectiologie de l'Association française d'urologie (CIAFU).

Resistance progression of the Neisseria gonorrhoeae to quinolones and the decreasing sensitivity to cephalosporin implicate to actualise the guidelines for managing urethritis. We present the guidelines from the committee of infectious diseases of the French Association of Urology to manage acute urethritis.

[Diagnosis, treatment and follow-up of community-acquired bacterial infections of the urinary system of men and women (acute cystitis and acute pyelonephritis) and of the genital system of men (acute prostatitis): general remarks]. / Généralités.

Urinary tract infections are frequent. The aim of these guidelines is to improve the management of urionary tract infections. Increasing antibiotic prescriptions may increase bacterial drug resistance. Asymptomatic bacteriuria, bacterial count, pyuria are defined and the clinical value of the bacterial culture and urinary dipstick test are discussed. The good antibiotic use depends on bacteriological, pharmaceutical, patient characteristics and economic findings which are precised in these guidelines.

[Acute cystitis]. / Cystites aiguës.

The management of uncomplicated lower urinary tract infections (UTI) implicate to look for risk factors and complications. Bacterial or radiological exams are not recommanded and short course of antibiotic is effective for treating uncomplicated UTI. Complicated UTI needs clinical, bacteriological and radiological exams, longer treatments are recommanded. Recurrent UTI definition is precised in these guidelines.

[Acute pyelonephritis]. / Pyélonéphrites aiguës.

The initial management of pyelonephritis needs to look for complicating factors. Ultrasound and X ray of the abdomen are able to rule out a urinary dilatation or a stone. The treatment is then surgical with renal drainage. Additional investigations such as a CT scan should be performed in patients with complicating factors or recurrence. In uncomplicated pyelonephritis a ambulatory treatment with 2 weeks of fluoroquinolones or cephalosporine Gr3 is sufficient. More severe cases should be admitted to a hospital and treated with initial cephalosporin Gr 3 plus aminoside for 3 to 6 weeks.

[Acute prostatitis]. / Prostatites aiguës.

A urinary infection in a febrile man is classiquely defined as a prostatitis. Investigation exams look for complicating factors or post voiding residual which should be drained. Antibiotic treatment should begin with a fluroquinolone or cephalosporin gr 3 for 3 to 6 weeks.

[In vitro activity of levofloxacin against Escherichia coli strains in acute pyelonephritis, in France in 2005]. / Activité in vitro de la lévofloxacine vis-à-vis de souches de Escherichia coli responsables de pyélonéphrites aiguës communautaires en France en 2005.

The objective of this study was to assess the in vitro activity of levofloxacin compared to other antibiotics against Escherichia coli strains isolated from female patients with acute pyelonephritis in 23 French hospitals in 2005. Minimal inhibitory concentrations (MICs) were determined in a central laboratory, by agar dilution method in compliance with the recommendations of the Comité de l'antibiogramme de la Société française de Microbiologie (CA-SFM). The percentages of susceptible strains were calculated according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) breakpoints approved by the CA-SFM (2005) for ciprofloxacin, levofloxacin, and ofloxacin and according to the CA-SFM breakpoints for the other antibiotics. Amongst the 231 strains collected, 46.3% of strains were isolated from urinary samples, 10.8% from blood culture, and 42.9% from both. Concerning fluoroquinolones, the percentages of susceptibility were 93.1%, 90.5%, and 92.7% for levofloxacin, ofloxacin, and ciprofloxacin, respectively. For the other antibiotics, a higher percentage of susceptibility was observed with ceftriaxone (99.6%) and amikacin (94.8%), whereas a lower percentage of susceptibility was observed with amoxiclav (68.8%), and cotrimoxazole (65.4%). Levofloxacin exhibited a good in vitro activity against E. coli strains isolated from acute pyelonephritis with 93.1% of susceptible strains.

Occurrence of qnrA-positive clinical isolates in French teaching hospitals during 2002-2005.

Bacteria harbouring the novel qnrA plasmid-mediated mechanism of quinolone resistance have been described in different countries, but the frequency of their occurrence has not been investigated. In total, 1,468 clinical isolates of Enterobacteriaceae with quinolone resistance or extended-spectrum beta-lactamase (ESBL) phenotypes were collected from eight teaching hospitals in France during 2002-2005 and screened for qnrA. Overall, 28 isolates (22 Enterobacter cloacae, three Klebsiella pneumoniae, one Citrobacter freundii, one Klebsiella oxytoca and one Proteus mirabilis) were positive for qnrA, representing 1.9% of all isolates, 3.3% of ESBL-producing isolates (22% of the E. cloacae isolates) and 0% of non-ESBL-producing isolates. The prevalence of qnrA among consecutive ESBL-producing isolates in 2004 from the eight hospitals was 2.8% (18/639). Of the qnrA-positive isolates, 100% were intermediately-resistant or resistant to nalidixic acid, and 75% to ciprofloxacin. Twenty-one of the 22 qnrA-positive E. cloacae isolates were obtained from two hospitals in the Paris area, and molecular typing and plasmid content analysis showed clonal relationships for five, three and two isolates, respectively. The qnrA genetic environment was similar to that of the In36 integron. The remaining two isolates had qnrA variants (30 and 29 nucleotide differences, respectively, compared with the original sequence) and an unknown genetic environment. The ESBL gene associated with qnrA was bla(SHV-12) in most of the isolates, but bla(PER-1) and bla(SHV-2a) were found in two isolates. In France, it appears that qnrA-positive isolates are predominantly E. cloacae isolates producing SHV-12, and may be associated with the dissemination of an In36-like integron.

European Committee on Antimicrobial Susceptibility Testing (EUCAST) Technical Notes on antimicrobial susceptibility testing.

The main objectives of the European Committee on Antimicrobial Susceptibility Testing (EUCAST) are to harmonise breakpoints for antimicrobial agents in Europe, and to act as the breakpoint committee for the European Medicines Agency (EMEA) during the registration of new antimicrobial agents. Detailed EUCAST procedures for harmonising and setting breakpoints for antimicrobial agents are available on the EUCAST website. Beginning with the current issue, a series of EUCAST Technical Notes will be published in CMI, based on the rationale documents produced by EUCAST for each of the antimicrobial agents studied, with the aim of highlighting important background information underlying decisions on breakpoints made by EUCAST.

Long-term efficacy of mupirocin in the prevention of infections with meticillin-resistant Staphylococcus aureus in a gastroenterology unit.

The long-term efficacy (55 months) of eradication of nasal carriage of meticillin-resistant Staphylococcus aureus (MRSA) by mupirocin was assessed for MRSA infections in a gastroenterology unit receiving patients for long hospital stays. In total, 2242 patients were included in the study; 92% had been hospitalized in another hospital before admission to the study department, 64% had chronic liver diseases (LD), 25% had miscellaneous medical conditions and 11% were admitted following gastroenterological surgery. Three consecutive periods were considered in the analysis. Nasal carriage at admission was similar in all three periods (10.9 vs 7.5 vs 8.6% in Periods 1, 2 and 3, respectively), while acquired nasal carriage decreased in the whole population (14.3 vs 16.2 vs 10.2% in Periods 1, 2 and 3, respectively, P=0.006) and in LD patients (15.8 vs 18.7 vs 11.9% in Periods 1, 2 and 3, respectively, P=0.018). The incidence of MRSA infections (N per total number of hospitalization-days) was 1.41 per 1000 in the year before initiation of eradication, 1.40 in Period 1, 0.74 in Period 2 and 0.59 in Period 3 (P=0.022). The incidence of MRSA infections among patients was 7.0% in Period 1, 3.7% in Period 2 and 3.1% in Period 3 in LD patients (P=0.0062). The corresponding figures were 5.5, 3.0 and 2.4% for the whole population (P=0.0024). The mortality caused by MRSA was 0.31, 0.19 and 0.13% (P=0.035) in Periods 1, 2 and 3, respectively. The numbers of resistant strains among those acquired during hospitalization were 12 in Period 1, four in Period 2 and six in Period 3. Long-term intranasal mupirocin treatment in MRSA carrier patients with long hospital stay is associated with a decrease in acquired carriage and MRSA infections, while resistance of the strains to mupirocin does not increase provided that colonized patients are only treated once.

Distribution and antimicrobial susceptibility of bacteria from adults with community-acquired pneumonia or complicated skin and soft tissue infections in France: the nationwide French PREMIUM study.

The empirical therapy of community-acquired pneumonia (CAP) and complicated skin and soft tissue infections (cSSTIs) must be based on updated bacterial distribution and susceptibility data. A nationwide study consecutively collected 1288 isolates from CAP (n=467) and cSSTIs (n=821) from 18 French hospitals between 2012 and 2013. The MIC values of commonly used antimicrobial agents, including ceftaroline, were determined. Bacterial distribution featured Pneumococcus, Haemophilus influenzae, and Staphylococcus aureus for CAPs and S. aureus, ß-hemolytic streptococci and Enterobacteriaceae for cSSTIs. Antimicrobial susceptibility testing indicated i) the sustained third-generation cephalosporins and levofloxacin activity against pneumococci and H. influenzae, ii) no methicillin-resistant Staphylococcus aureus emergence among respiratory pathogens, iii) the high in vitro activity of ceftaroline against staphylococci from cSSTIs (98.7% susceptibility), and iv) the worrisome decreasing fluoroquinolone and third-generation cephalosporin susceptibilities among Enterobacteriaceae. This laboratory-based survey depicts a contrasting situation and supports the scoring of patients for the resistant pathogen risk before empirical therapy.

Impact of Helicobacter pylori resistance to clarithromycin on the efficacy of the omeprazole-amoxicillin-clarithromycin therapy.

BACKGROUND: Helicobacter pylori resistance to clarithromycin is relatively frequent in France and is assumed to be the main cause of failure of the proton pump inhibitor-amoxicillin-clarithromycin (proton pump inhibitor-AC) therapy, which is the first-line regimen in France. AIM: To determine the respective effects of clarithromycin primary and secondary resistances on efficacy of the proton pump inhibitor-AC regimen and to determine whether failures are associated with persistence of the same strain or with emergence of a new one. METHODS: A total of 123 H. pylori-infected patients were treated for 7 days with omeprazole 20 mg b.d., amoxicillin 1 g b.d., and clarithromycin 500 mg b.d. Eradication was assessed by breath test in 102 patients. Minimal inhibitory concentrations of clarithromycin were determined by E-test. Strain genotyping was performed by random amplified polymorphic DNA. RESULTS: The pre-treatment and post-treatment prevalences of clarithromycin resistance were 19% (23 out of 123) and 69% (nine out of 13), respectively. The rates of eradication were 68% (69 out of 102), 79% (67 out of 85), and 12% (two out of 17) for all, susceptible and resistant strains, respectively. The post-treatment isolate was available for six patients with a susceptible pre-treatment isolate and a persistent infection. Resistance emerged in two patients and was associated with persistence of the pre-treatment strain in one and with selection of a new strain in the other. CONCLUSIONS: In our hospital, failures of the proton pump inhibitor-AC therapy are related to both clarithromycin primary and secondary resistances, but the emergence of secondary resistance does not explain all of the failures in the initial clarithromycin-susceptible group. In that group a new strain can emerge after failure.

Analysis of the mutations involved in fluoroquinolone resistance of in vivo and in vitro mutants of Escherichia coli.

We searched for the mutations involved in high-level fluoroquinolone resistance (ciprofloxacin MIC > or = 8 microg/ml) of 11 clinical isolates of Escherichia coli. trans-Complementation tests with the wild-type gyrA and parC genes were positive for all strains whereas negative results were observed with the wild-type gyrB and parE genes. By PCR and sequencing, two mutations in gyrA, leading to Ser-83 --> Leu and Asp-87 --> Asn (7) or Gly (2) or Tyr (1) changes, were found in 10 strains, the eleventh presenting only the Ser-83 --> Leu change. In addition, all strains carried one change in ParC: Ser-80 --> Ile (8) or Arg (2); Glu-84 --> Lys (1). We described a novel and simple method permitting detection of the mutations in parC at codon 80, PCR-RFLP with HaeII. In vitro mutants, selected with ciprofloxacin in three successive steps were also studied. The first-step mutants were complemented by pJSW101 (gyrA+) but not by pEN260 (parC+), whereas the second-step and third-step mutants were complemented by both plasmids. Mutations occurred in the following order: (i) gyrA at codon 83 (Ser to Leu change), (ii) parC at codon 80 (Ser to Ile change), and (iii) gyrA at codon 87 (Asp to Asn change). Thus, these sequential mutations appear to be frequently involved in high-level fluoroquinolone resistance of E. coli.

Quinolone resistance mutations in the gyrA gene of clinical isolates of Salmonella.

S. typhimurium AlhR, S. enteritidis OulR, and S. hadar GueR resistant to fluoroquinolones (QR), ciprofloxacin MICs, 0.25 to 1 microgram/ml; norfloxacin MICs, 0.5 to 4 micrograms/ml; nalidixic acid MIC, 256 micrograms/ml were isolated from urinary tract infections (AlhR and OulR) during FQ therapy in immunocompromised patients infected by the parent FQ-susceptible strains (AlhS and OulS) (ciprofloxacin MICs, 0.032-0.063; norfloxacin MICs, 0.125-0.25; nalidixic acid MICs, 4-8) or from intestinal infection (GueR). Transformation of AlhR, OulR, and GueR by plasmid pJSW101 carrying the wild-type gyrA gene of Escherichia coli resulted in complementation (nalidixic acid MICs, 4 to 8), proving that these strains had a gyrA mutation. A 800-bp fragment of gyrA from the five strains was amplified by PCR. Direct DNA sequencing of 252-bp region of this fragment identified a single point mutation leading to a substitution Ser-83 to Tyr in AlhR and to a substitution Ser-83 to Phe in OulR and in GueR. These results emphasize the potential risk of selection of FQ-resistant Salmonella during FQ therapy in immunocompromised patients and suggest that these strains differ from the parent strains at least by one mutation in the gyrA gene. They also confirm the role of substitutions in position 83 of gyrA in FQ-resistant clinical isolates of Salmonella.

Single or double mutational alterations of gyrA associated with fluoroquinolone resistance in Campylobacter jejuni and Campylobacter coli.

We looked for the presence of gyrA mutations in seven fluoroquinolone-resistant French clinical isolates of Campylobacter jejuni and Campylobacter coli. Three of the five isolates of C. jejuni and the two isolates of C. coli had high-level resistance to nalidixic acid (MICs 128-256 microg/ml) and ciprofloxacin (MICs 32 microg/ml). A gyrA mutation was found in all these isolates leading to the following substitutions: Thr86-Ile in four cases and Asp90-Tyr for one C. coli strain. One isolate had high-level resistance to nalidixic acid (MIC 64 microg/ml) but low-level resistance to ciprofloxacin (MIC 2 microg/ml) and also carried a gyrA mutation leading to a Thr86-Ala substitution. The last isolate of C. jejuni studied displayed an atypical resistance phenotype: It was resistant to high levels of ciprofloxacin (MIC 64 microg/ml) but remained fully susceptible to nalidixic acid (MIC 2 microg/ml). This phenotype was not explained by the presence of peculiar mutations in gyrA or gyrB. It carried a gyrA mutation leading to a Thr86-Ile substitution and was devoid of gyrB mutation. Despite numerous attempts with various degenerate oligonucleotide primers deduced from conserved regions of known parC genes, we were unable to amplify a corresponding sequence in C. jejuni or C. coli. First-step and second-step in vitro mutants, derived from reference strain C. coli ATCC 33559 with ciprofloxacin or moxifloxacin as selecting agents, were found to carry one and two mutations in gyrA, respectively. In contrast with the results obtained with clinical isolates, a variety of gyrA mutations were obtained in vitro.

European Glycopeptide Susceptibility Survey of gram-positive bacteria for 1995. European Glycopeptide Resistance Survey Study Group.

In the European Glycopeptide Susceptibility Survey 7078 Gram-positive isolates collected in 1995 from 70 centers in 9 countries of Western Europe were examined, using a standardized, quantitative susceptibility testing method. Of the 7078 isolates, 6824 (96.4%) were tested by the national coordinating centers. Teicoplanin (mode MIC 0.5 microgram/mL) was generally twice as active as vancomycin (mode MIC 1 microgram/mL) against Staphylococcus aureus (n = 2852). All isolates were susceptible to vancomycin (MIC < or = 4 micrograms/mL) and all but four to teicoplanin (MIC < or = 8 micrograms/mL); these four isolates were of intermediate susceptibility (MIC 16 micrograms/mL). With coagulase-negative staphylococci (n = 1444), the distribution of MIC of teicoplanin was wider than for vancomycin. Two and two-tenths percent of coagulase-negative staphylococci excluding Staphylococcus haemolyticus required 16 micrograms/mL teicoplanin for inhibition (intermediate) and 0.4% > or = 32 micrograms/mL (resistant). Among isolates of S. haemolyticus, 4.4% were of intermediate susceptibility (MIC 16 micrograms/mL) and 3.3% were resistant (MIC > or = 32 micrograms/mL) to teicoplanin. However, this species represented only 6.3% of the isolates of coagulase-negative Staphylococcus spp. Generally, teicoplanin (mode MIC < or = 0.12 microgram/mL) was four to eight times more active than vancomycin (mode MIC < or = 0.5 microgram/mL) against the 770 streptococcal isolates. Glycopeptide-susceptible Enterococcus spp. (n = 1695) were generally four times more susceptible to teicoplanin (mode MIC 0.25 microgram/mL) than to vancomycin (mode MIC 1 microgram/mL). Combined vancomycin and teicoplanin (VanA phenotype) resistance was observed more frequently (9.3%) in isolates of Enterococcus faecium than in Enterococcus faecalis (0.8%). Four isolates of unspeciated enterococci (1.4%) also expressed this resistance phenotype. Four isolates of E. faecium and four of E. faecalis expressed the VanB-type (low-level, vancomycin only) resistance. Spain was the only country not to submit resistant E. faecium strains while resistant E. faecalis isolates came only from Spain and Italy.

Co-detection of Helicobacter pylori and of its resistance to clarithromycin by PCR.

Our aim was to develop a rapid molecular test based on polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and making it possible to detect Helicobacter pylori directly from gastric biopsy samples, and to test its susceptibility to clarithromycin. A 629-bp fragment of the 23S rRNA gene of H. pylori was amplified by PCR and the mutations responsible for clarithromycin resistance were detected with Bsa1 and Bbs1 restriction endonucleases. Thirty-five gastric samples were tested in parallel by standard microbiologic methods (culture and clarithromycin susceptibility testing with E-test strips) and by PCR-RFLP. The 10 culture-negative samples were also PCR-negative. Sixteen out of the 25 culture-positive samples (64%) were PCR-positive. RFLP analysis could be done in 12 cases and the results were in agreement with those of the E-test: susceptibility in five cases, resistance in seven (six A2144G mutations and one A2143G mutation).

What technique should be used for routine detection and quantification of HBV DNA in clinical samples?

Detection of hepatitis B virus (HBV) DNA in serum allows monitoring of HBV replication and assessing responses to antiviral treatment. HBV DNA quantification measures virus replication and can be used as a prognosis indicator of liver disease and an index of response to antiviral drugs. The aim of this study was to compare the performances of three HBV DNA detection and/or quantification techniques for assessing HBV replication. Three hundred unselected sera with a request for HBV DNA detection and quantification were tested with a molecular hybridisation technique without amplification (Digene Hybrid-Capture, Murex Diagnostics Ltd), a signal amplification assay based on branched DNA technology (Quantiplex HBV DNA, Chiron diagnostics), and an 'in-house' qualitative, non quantitative target amplification assay based on the polymerase chain reaction (PCR) with primers located in the S gene of the HBV genome. Hybrid-capture and branched DNA gave concordant results in 278 cases (93%). In the 128 samples positive by both assays, DNA titres in pg/ml were related significantly (r = 0.70, P < 0.0001). but branched DNA titres increased more rapidly than hybrid-capture titres when the amount of HBV DNA in the sample increased. Twenty-two sera (7%) were negative by hybrid-capture, but positive in branched DNA (detection rate gain: 15%). In these 22 patients, DNA titres were low, HBsAg was present in all instances and alanine aminotransferase activity was elevated in 18 patients (82%); HBeAg was present in seven patients (32%) and anti-HBe antibodies in 18 patients (82%); liver biopsy, undertaken in 18 patients, revealed chronic active hepatitis in all instances, associated with cirrhosis in eight cases. Qualitative, non-quantitative HBV DNA PCR was positive in 75 (50%) of the 150 hybrid-capture-negative, branched DNA-negative samples, including a significant proportion of patients without evidence of ongoing HBV-related liver disease. The results show that in general, the branched DNA assay detects HBV DNA in more patients than hybrid-capture and that this improved detection rate is relevant clinically and genome equivalents/ml are preferred to pg/ml to quantify HBV DNA in clinical specimens and finally qualitative, non-quantitative polymerase chain reaction can detect HBV DNA in patients without evidence of active HBV-related liver disease. This study emphasizes the need for more sensitive, university standardised quantitative HBV DNA assays and for the definition of clinically relevant cutoffs with these assays.