Improved quality of care for patients infected or colonised with ESBL-producing Enterobacteriaceae in a French teaching hospital: impact of an interventional prospective study and development of specific tools.

The increasing incidence of ESBL-producing Enterobacteriaceae (ESBL-E) in France prompted the publication of national recommendations in 2010. Based on these, we developed a toolkit and a warning system to optimise management of ESBL-E infected or colonised patients in both community and hospital settings. The impact of this initiative on quality of care was assessed in a teaching hospital. The ESBL toolkit was developed in 2011 during multidisciplinary meetings involving a regional network of hospital, private clinic and laboratory staff in Southeastern France. It includes antibiotic treatment protocols, a check list, mail templates and a patient information sheet focusing on infection control. Upon identification of ESBL-E, the warning system involves alerting the attending physician and the infectious disease (ID) advisor, with immediate, advice-based implementation of the toolkit. The procedure and toolkit were tested in our teaching hospital. Patient management was compared before and after implementation of the toolkit over two 3-month periods (July-October 2010 and 2012). Implementation of the ESBL-E warning system and ESBL-E toolkit was tested for 87 patients in 2010 and 92 patients in 2012, resulting in improved patient management: expert advice sought and followed (16 vs 97%), information provided to the patient's general practitioner (18 vs 63%) and coding of the condition in the patient's medical file (17 vs 59%), respectively. Our multidisciplinary strategy improved quality of care for in-patients infected or colonised with ESBL-E, increasing compliance with national recommendations.

Role of DNA gyrase and topoisomerase IV mutations in fluoroquinolone resistance of Capnocytophaga spp. clinical isolates and laboratory mutants.

Objectives: Capnocytophaga spp. are often reported to cause bacteraemia and extra-oral infections and are characterized by their significant contribution to resistance to ß-lactam and macrolide-lincosamide-streptogramin antibiotics in the human oral microbiota. The implication of mutations in the quinolone resistance-determining region (QRDR) of DNA gyrase A and B ( gyrA and gyrB ) and topoisomerase IV ( parC and parE ) of fluoroquinolone (FQ)-resistant Capnocytophaga spp., hitherto unknown, was explored in this study. Methods: Two reference strains ( Capnocytophaga gingivalis ATCC 33624 and Capnocytophaga sputigena ATCC 33612) and four Capnocytophaga spp. isolated from clinical samples were studied. Nine in vitro FQ-resistant mutants, derived from two reference strains and one FQ-susceptible clinical isolate, were selected by successive inoculations onto medium containing levofloxacin. MICs of ofloxacin, norfloxacin, ciprofloxacin, levofloxacin and moxifloxacin were determined. The presumed QRDRs of GyrA, GyrB, ParC and ParE from Capnocytophaga spp. were determined by sequence homology to Bacteroides fragilis and Escherichia coli . PCR primers were designed to amplify the presumed QRDR genetic region of Capnocytophaga spp. and sequence analyses were performed using the BLAST program at the National Center for Biotechnology Information. Results and conclusions: gyrA mutations leading to a substitution from amino acid position 80 to 86 were systematically detected in Capnocytophaga spp. with ciprofloxacin MIC >1 mg/L and considered as the primary target of FQs. No mutational alteration in the QRDR of gyrB was detected. Other mutations in parC and parE led to spontaneous amino acid substitutions of DNA topoisomerase IV subunit B with no alteration in FQ susceptibility.

Multidrug-resistant oral Capnocytophaga gingivalis responsible for an acute exacerbation of chronic obstructive pulmonary disease: Case report and literature review.

INTRODUCTION: Capnocytophaga genus was recently known to highly contribute to the beta-lactam (BL) and macrolide-lincosamide-streptogramin (MLS) resistance gene reservoir in the oral microbiota (BL: blaCSP-1 and blaCfxA; MLS: erm(F) and erm(C)). But fluoroquinolone (FQ) resistance remains uncommon in literature, without available data on resistance mechanisms. CASE REPORT: For the first time, a case of acute exacerbation of chronic obstructive pulmonary disease (COPD) was described in a 78-year-old immunocompetent patient due to a multidrug-resistant Capnocytophaga gingivalis isolate with significant microbiological finding. C.gingivalis acquired resistance to third generation cephalosporins (blaCfxA3 gene), MLS (erm(F) gene), and fluoroquinolones. Genetics of the resistance, unknown as regards fluoroquinolone, was investigated and a substitution in QRDR of GyrA was described (Gly80Asn substitution) for the first time in the Capnocytophaga genus. LITERATURE REVIEW: A comprehensive literature review of Capnocytophaga spp. extra-oral infection was conducted. Including the present report, on 43 cases, 7 isolates were BL-resistant (17%), 4 isolates were MLS-resistant (9.5%) and 4 isolates were FQ-resistant (9.5%). The studied clinical isolate of C.gingivalis was the only one to combine resistance to the three groups of antibiotics BL, MLS and FQ. Four cases of Capnocytophaga lung infection were reported, including three infections involving C. gingivalis (two FQ resistant) and one involving C. sputigena. CONCLUSION: This multidrug-resistant C. gingivalis isolate illustrated the role of oral flora as a reservoir of antibiotic resistance and its contribution to the limitation of effective antibiotics in severe respiratory infections.

High prevalence of ß-lactam and macrolide resistance genes in human oral Capnocytophaga species.

OBJECTIVES: To determine macrolide-lincosamide-streptogramin (MLS) resistance determinants in the Capnocytophaga genus and to describe the prevalence of ß-lactam resistance genes in human oral Capnocytophaga species. METHODS: Forty-eight Capnocytophaga isolates identified by analysis of 16S rRNA sequences were isolated from subgingival samples from 14 haematology patients (HPs), 11 periodontitis patients (PPs) and 17 healthy volunteers (HVs). MICs of ß-lactam and MLS antibiotics were obtained for all isolates. blaCfxA, blaCSP-1 (encoding a new class A ß-lactamase) and MLS resistance genes [erm(F), erm(B), erm(Q), erm(D), erm(C) and erm(A)] were evaluated using specific PCR and sequencing. RESULTS: In HVs, which had the lowest prevalence of ß-lactamase-producing isolates in comparison with the other groups (16%; P < 0.001), Capnocytophaga ochracea was the prominent species (68%; P < 0.03). In PPs, which had a high prevalence of ß-lactamase-positive isolates (82%; P < 0.001), Capnocytophaga sputigena was more frequently identified (64%; P < 0.03). In HPs, 50% of isolates were ß-lactamase-positive. The more rarely identified species (15%) Capnocytophaga gingivalis, Capnocytophaga granulosa and Capnocytophaga leadbetteri were isolated only from PPs and/or HPs. All ß-lactam-resistant isolates (44%) were PCR-positive for blaCfxA (31%) or blaCSP-1 (12.5%). Interestingly, blaCSP-1 was identified only in a subgroup of the C. sputigena species. Twenty-nine percent of isolates were MLS resistant independently of species identification, ß-lactamase production or patient group. The MLS-resistant isolates carried the erm(F) or erm(C) gene (93% and 7%, respectively), previously unknown in the Capnocytophaga genus. CONCLUSIONS: Our findings illustrate that Capnocytophaga species are important contributors to the ß-lactam and MLS resistance gene reservoir in the oral microbiome.

National multidrug-resistant bacteria (MDRB) surveillance in France through the RAISIN network: a 9 year experience.

BACKGROUND: In the mid-1990s, the prevalence rate of multidrug-resistant bacteria (MDRB) in French hospitals was high and control of MDRB spread then became a major priority in the national infection control programme (ICP). METHODS: To evaluate the impact of the ICP, a national coordination of MDRB surveillance was set up in 2002. Data were collected 3 months a year in healthcare facilities (HCFs) on a voluntary basis. All clinical specimens of methicillin-resistant Staphylococcus aureus (MRSA) and extended-spectrum ß-lactamase-producing Enterobacteriaceae (ESBLE) were prospectively included. Incidences per 1000 patient days (PDs) were calculated and trends in incidence from 2003 to 2010 were assessed. RESULTS: Participation in the surveillance increased from 478 HCFs in 2002 to 933 in 2010. In 2010, MRSA incidence was 0.40/1000 PDs: 1.14 in intensive care units (ICUs), 0.48 in acute care facilities (ACFs) and 0.27 in rehabilitation and long-term care facilities (RLTCFs). ESBLE incidence was 0.39/1000 PDs: 1.63 in ICUs, 0.46 in ACFs and 0.23 in RLTCFs. MRSA incidence significantly decreased from 0.72/1000 PDs in 2003 to 0.41/1000 PDs in 2010 (P<10(-3)); in contrast, ESBLE incidence significantly increased from 0.17/1000 PDs to 0.48/1000 PDs (P<10(-3)). The most prevalent ESBLE were Enterobacter aerogenes (34%) and Escherichia coli (25%) in 2003 and E. coli (60%) and Klebsiella pneumoniae (18%) in 2010. CONCLUSION: These results demonstrate the positive impact of the national ICP on MRSA rates. In contrast, ESBLE incidence, especially ESBL-producing E. coli, is increasing dramatically and represents a serious threat for hospitals and for the community that deserves specific control actions.

Genetic and biochemical characterization of TRU-1, the endogenous class C beta-lactamase from Aeromonas enteropelogenes.

Aeromonas enteropelogenes (formerly A. tructi) was described to be an ampicillin-susceptible and cephalothin-resistant Aeromonas species, which suggests the production of a cephalosporinase. Strain ATCC 49803 was susceptible to amoxicillin, cefotaxime, and imipenem but resistant to cefazolin (MICs of 2, 0.032, 0.125, and >256 microg/ml, respectively) and produced an inducible beta-lactamase. Cefotaxime-resistant mutants (MIC, 32 microg/ml) that showed constitutive beta-lactamase production could be selected in vitro. The gene coding for the cephalosporinase of A. enteropelogenes ATCC 49803 was cloned, and its biochemical properties were investigated. Escherichia coli transformants showing resistance to various beta-lactams carried a 3.5-kb plasmid insert whose sequence revealed a 1,146-bp open reading frame (ORF) encoding a class C beta-lactamase, named TRU-1, showing the highest identity scores with A. punctata CAV-1 (75%), A. salmonicida AmpC (75%), and A. hydrophila CepH (71%). The bla(TRU-1) locus includes open reading frames (ORFs) showing significant homology with genes found in the genomes of other Aeromonas species, although it exhibits a different organization, as reflected by the presence of additional ORFs located downstream of the beta-lactamase gene in the A. hydrophila and A. salmonicida genomes. Specific PCR assays were negative for cphA-like and bla(OXA-12)-like genes in three A. enteropelogenes ATCC strains. Purified TRU-1 showed a broad substrate profile, efficiently hydrolyzing benzylpenicillin, cephalothin, cefoxitin, and, although with significantly lower turnover rates, oxyiminocephalosporins. Cephaloridine and cefepime were poorly recognized by the enzyme, as reflected by the high K(m) values observed with these substrates. Thus far, A. enteropelogenes represents the only known example of an Aeromonas species that produces only one beta-lactamase belonging to molecular class C.

Antimicrobial resistance data on 16,756 Streptococcus pneumoniae isolates in 1999: A Pan-Regional Multicenter Surveillance Study in France.

The rising prevalence of antibiotic-resistant Streptococcus pneumoniae is a phenomenon observed to different degrees around the world. The present national surveillance study report analyzes a total of 16,756 strains of S. pneumoniae collected across France in 1999. The overall prevalence of S. pneumoniae with decreased susceptibility to penicillin was 44%, to amoxicillin 26%, and to cefotaxime 17%. The proportion of high-level resistant strains to penicillin (MIC > 1 mg/L), amoxicillin and cefotaxime (MIC > 2 mg/L) remained low: 12.3%, 1.8%, and 0.4% respectively. Prevalence of resistance to other antibiotics was high: 53% to erythromycin, 41.7% to cotrimoxazole, 31.8% to tetracycline, and 24.6% to chloramphenicol. Prevalence of penicillin-resistant S. pneumoniae varied according to subject age and specimen source. It was higher in children (52.7%) than in adults (39.8%) and higher in strains isolated from middle ear fluid (63.6%) than from blood cultures (41.8%) in children. S. pneumoniae resistant to other antibiotics were more common in children than in adults, although figures showed geographical variations. Comparison with a previous study realized in 1997 in the same regions confirms a rising trend in the prevalence of resistant bacteria. Therefore, we conclude that prevalence of antibiotic-resistant S. pneumoniae in 1999 continued to rise in France, although strains with high-level resistance to penicillin remained stable.

Sequence analysis and biochemical characterisation of chromosomal CAV-1 (Aeromonas caviae), the parental cephalosporinase of plasmid-mediated AmpC 'FOX' cluster.

Aeromonas caviae CIP 74.32 was resistant to amoxicillin, ticarcillin and cephalothin, and susceptible to cefoxitin, cefotaxime, ceftazidime, aztreonam and imipenem. This strain produced a cephalosporinase (pI 7.2) and an oxacillinase (pI 8.5). The cephalosporinase gene cav-1 was cloned and sequenced. Unlike A. caviae donor, Escherichia coli pNCE50 transformant producing CAV-1 beta-lactamase was resistant to cefoxitin. The deduced protein sequence CAV-1 contained 382 amino acids, and shared >96% homology with FOX-1 to FOX-5 cephalosporinase. CAV-1 presented only two amino acid substitutions (Thr270Ser and Arg271Ala) with FOX-1. CAV-1 is the chromosomal putative ancestor of the FOX family, a cluster of class C/group 1 plasmidic cephalosporinases spreading in Klebsiella and E. coli clinical isolates via conjugative plasmids.

[The buccal-dental health of drug addicts treated in the University hospital centre in Nice]. / Etat de santé bucco-dentaire des toxicomanes suivis au CHU de Nice.

OBJECTIVE: Drug-addicts often suffer from rapidly progressive extensive dental decay. The objectives of this study were to determine the impact of illicit drugs on buccal and dental health and the use of illicit drugs for toothache. METHODS: Two groups of intravenous and non-intravenous drug-addicts were compared with two control groups of age matched non-addicted subjects. During a routine dental examination, medical and addictive history, periodontal and dental health and dental complaints were recorded. RESULTS: This study showed that intravenous heroin was responsible for rapidly progressive dental decay, even in four drug-addicts with satisfactory dental hygiene. Intravenous heroin users (14 women, 38 men, mean age 35) had a mean number of 10 missing and 10 decayed teeth, 6 of them to be extracted, and needed two dentures with 8 teeth each. Their masticatory function (45%) and smile did not permit normal alimentation or social life. Non-intravenous drug users (9 women, 29 men, mean age 26) had a mean number of one missing tooth and 4 decayed teeth to be treated. When compared to control groups, drug users of both categories exhibited more decayed teeth, reduced masticatory function and a lower periodontal health correlated with inadequate dental hygiene. Finally, 52% of heroin users and 21% of other illicit drug users admitted the use of illicit drugs as analgesics for toothache. CONCLUSION: The management of toothache should be proposed in the cessation protocols and dentures provided to intravenous drug-addicts, before any attempt at social reinsertion.

[Escherichia coli bacteremic urinary tract infections. Clinical aspects and pronostic factors]. / Bactériémies à Escherichia coli d'origine urinaire. Caractéristiques cliniques et facteurs pronostiques.

OBJECTIVE: To describe patients with bacteremia from urinary tract infection caused by Escherichia coli and investigate risk factors for mortality. METHOD: Retrospectively study of the files of patients with bacteremic urinary tract infection caused by E. coli and admitted via the Emergency Department of the University Hospital in Nice between 01/01/1997 and 12/31/2000. Inclusion criteria included at least one blood and urine culture positive for E. coli during the first 48 hours and age above 15 years. RESULTS: There were 118 patients in the population (71% female). Mean age was 73 years and median age 79 years. The majority of patients (90%) were hospitalized in a medical department. Initially the clinical picture was sepsis in 80% of patients, severe sepsis in 15% and septic shock in 5%. Lethality was 16%. In 40% of cases death occurred within the first 48 hours. Risk factors for mortality in multivariate analysis were initially severe clinical status and male sex. CONCLUSION: The population was aged and mortality was high for an infection presumed to be relatively benign. Age was not a risk factor for mortality, contrary to male gender.

Indicadores das infecções associadas aos cuidados e gestão do risco infeccioso: a experiência francesa / Indicators of the infections associated with care and infectious risk management: the french experience / Indicadores de las infecciones asociadas a los cuidados y gestión del riesgo infeccioso: la experiencia francesa

Este artigo apresenta como são organizadas as ações contra as Infecções Associadas aos Cuidados na França e discute os indicadores associados aos cuidados e gestão do risco infeccioso. Trata da reorganização e modernização do sistema de saúde francês no que se refere à política de melhorias contínuas da qualidade, segurança do cuidado, dos dispositivos, funcionamento e estruturas de controle das Infecções Associadas aos Cuidados. A gestão do risco infeccioso está associada à certificação obrigatória, sendo realizada em rede articulada e conforme diretrizes da Política e do Programa de Prevenção das Infecções Associadas aos Cuidados. Os indicadores fazem parte do painel público de infecções hospitalares, podendo o usuário ter acesso pelo site www.scopesante.fr. Os resultados revelam que, apesar dessa política, a França, entre os países da comunidade europeia, é o quarto país que mais utiliza antibióticos em nível hospitalar. A vigilância do consumo de antibióticos nos estabelecimentos de saúde de todo o país é uma estratégia para reduzir o seu uso. Além disso, são necessárias ações específicas para aprender a melhor prescrever antibióticos. Os resultados, com base em objetivos quantificados e indicadores, permitem verificar quais e onde os esforços precisam aumentar.

This article presents how to organize actions against infections associated to care in France and discuss the indicators associated with the care and the management of infectious risk. It talks about the reorganization and the modernization of the French health system with regards to the policy of continuous improvement of quality, safety of care, of devices, of operation and control structures of Care Associated Infections. Infectious Risk management is associated with the mandatory certification, and carried out in an articulated network and according to guidelines of the Policy and Prevention Program of Care Associated Infections. The indicators are part of the public panel of hospital infections, allowing access to the user on the site www.scopesante.fr. The results show that, despite the policy among the countries of the European Community, France is the fourth country using more antibiotics in hospitals. The control of antibiotic consumption in health facilities across the country is a strategy to reduce its use. In addition, specific actions are needed to learn how to best prescribe antibiotics. The results based on quantified targets and indicators evince what and where efforts need to increase.

Este artículo presenta como se organizan las acciones contra las Infecciones Asociadas a los Cuidados en Francia y discute los indicadores asociados a los cuidados y la gestión del riesgo infeccioso. Trata de la reorganización y modernización del sistema de salud francés en lo que se refiere a la política de mejorías contínuas de la calidad, seguridad del cuidado, de los dispositivos, del funcionamiento y de las estructuras de control de las Infecciones Asociadas a los Cuidados. La gestión del riesgo infeccioso está asociada a la certificación obligatoria, siendo realizada en red articulada y según directivas de la Política y del Programa de Prevención de las Infecciones Asociadas a los Cuidados. Los indicadores hacen parte del panel público de infecciones hospitalarias, a que el usuario puede acceso a través del site www.scopesante.fr. Los resultados revelan que, a pesar de esa política, la Francia, entre los países de la Comunidad Europea, es el cuarto país que más utiliza antibióticos en nivel hospitalario. La vigilancia del consumo de antibióticos en los centros sanitarios de todo el país es una estrategia para reducir su uso. Además, son necesarias acciones específicas para aprender a prescribir mejor los antibióticos. Los resultados, con base en objetivos cuantificados e indicadores, permiten verificar cuales y donde los esfuerzos son necesarios.

Epidemiology and antimicrobial resistance of Streptococcus pneumoniae in France in 2007: data from the pneumococcus surveillance network.

Antimicrobial resistance of Streptococcus pneumoniae in France is closely monitored by the pneumococcus surveillance network, founded in 1995, which collects data from regional observatories (Observatoire Régionaux du Pneumocoque [ORP]). In 2007, 23 ORPs analyzed the antibiotic susceptibility of 5,302 isolates of S. pneumoniae recovered in France from cerebrospinal fluid, blood, middle ear fluid, and pleural fluid, as well as from adult respiratory samples. The study showed that 38.2% of the strains were nonsusceptible to penicillin, 19.3% nonsusceptible to amoxicillin, and 10.5% nonsusceptible to cefotaxime. The percentage of pneumococcus nonsusceptible to penicillin varied according to both the sample and the age of the patient (child/adult): blood (27.8%/32.5%), cerebrospinal fluid (33.7%/34.6%), middle ear fluid (60.2%/27.5%), and pleural fluid (50.0%/31.0%). Between 2003 and 2007, the frequency of penicillin resistance in invasive pneumococcal disease gradually decreased from 46.4% to 29.0% in children and from 43.8% to 32.7% in adults. This decrease coincided with the introduction of a seven-valent pneumococcal conjugate vaccine into immunization programs and with a general reduction in levels of antibiotic consumption in France.

Serotype distribution and antibiotic resistance of Streptococcus pneumoniae strains isolated from adults in France: evolution between 2001 and 2003.

Antibiotic-resistant Streptococcus pneumoniae (Sp) are described around the world. The present national surveillance study report analyzes more than 6000 Sp strains, isolated from adults across France in 2001 and 2003, from blood cultures (3086 in 2001 and 3164 in 2003), cerebrospinal fluid (respectively, 238 and 240), or middle ear fluid (respectively, 110 and 100). The proportion of isolates with reduced susceptibility to penicillin fell significantly between 2001 and 2003 from 46.5% to 43.9%. The proportion of high-level resistant strains to penicillin minimal inhibitory concentrations (MIC > 1 mg/L), amoxicillin, and cefotaxime (MIC > 2 mg/L) slightly decreased but remained low: 10.6%, 1.2%, and 0.2% in 2003. Resistance to other antibiotics (erythromycin, cotrimoxazole, tetracycline, and chloramphenicol) also decreased. Decrease in prevalence of penicillin-resistant Sp varied according to specimen source. The proportion of penicillin nonsusceptible pneumococci decreased in blood cultures and middle ear fluids between 2001 and 2003 but increased in cerebrospinal fluid (43.4% and 46.5%, respectively). Serotypes covered by the heptavalent vaccine accounted for 42.4% of all isolates recovered in 2001 and 46.1% in 2003. Prevalence of antibiotic-resistant Sp decreased in 2003 in France.

["Antibiotics only when necessary" campaign in the Alpes-Maritimes district: no negative impact on invasive infections in children in the community 1998-2003]. / Campagne "Antibios quand il faut" dans les Alpes-Maritimes: absence d'impact sur l'incidence des infections invasives dues aux bactéries respiratoires communautaires chez l'enfant, 1998-2003.

OBJECTIVES: A multidisciplinary group (for the study and prevention of infections in children, GEPIE) conducted a local public health campaign to improve antibiotic prescribing in pediatric outpatient care in the Alpes-Maritimes district. This campaign included peer-conducted academic-detailing (educational outreach) visits in 2000 and 2003. Practitioners often report during both focus groups and the GEPIE visits that they prescribe antibiotics because of a fear of subsequent complications. This study therefore sought to explore the trends in invasive diseases related to bacterial respiratory pathogens. METHODS: A retrospective survey examined the incidence in the district of invasive infections due to Streptococcus pneumoniae, Haemophilus influenzae, Streptococcus pyogenes, and Neisseria meningitidis from 1998 through 2003 among children aged 1 month to 15 years. All local laboratories were asked for all reports of blood cultures (as well as spinal, synovial, pericardiac, peritoneal, and pleural fluid cultures) positive for these bacteria in the study population over the study period. Resistance and serotype data were recorded. RESULTS: Over the 6-year study period, there were 113 cases of invasive infections: 64% due to S. pneumoniae, 26% to N. meningitidis, 6% to H. influenzae, and 4% to S. pyogenes. There was no statistically significant difference in the annual average incidence rate before and during the campaign. The overall incidence rate was 11.2 cases per 100,000; meningitis accounted for 4.2, and invasive pneumococcal diseases 7.2 (42.7 per 100,000 children younger than 2 years). Pneumococcal resistance rates to penicillin and erythromycin did not change significantly. DISCUSSION: Two local cross-sectional studies at daycare centers in 1999 and 2002 showed a 9% reduction in the proportion of children who received antibiotics. The stability of invasive infection in children and of bacterial resistance during the campaign is reassuring. The campaign's risk/benefit ratio appears positive. CONCLUSION: These results encourage continuing promotion of rational antibiotic prescribing and ought to allay practitioners' fears about the consequences of prescribing antibiotics less often.

Genetic and biochemical characterization of FUS-1 (OXA-85), a narrow-spectrum class D beta-lactamase from Fusobacterium nucleatum subsp. polymorphum.

Previous studies have reported beta-lactamase-mediated penicillin resistance in Fusobacterium nucleatum, but no beta-lactamase gene has yet been identified in this species. An F. nucleatum subsp. polymorphum strain resistant to penicillin and amoxicillin was isolated from a human periodontitis sample. DNA cloning and sequencing revealed a 765-bp open reading frame encoding a new class D beta-lactamase named FUS-1 (OXA-85). A recombinant Escherichia coli strain carrying the bla(FUS-1) gene exhibited resistance to amoxicillin with a moderate decrease in the MICs with clavulanic acid. The bla(FUS-1) gene was found in two additional clonally unrelated F. nucleatum subsp. polymorphum isolates. It was located on the chromosome in a peculiar genetic environment where a gene encoding a putative transposase-like protein is found, suggesting a possible acquisition of this class D beta-lactamase gene. The FUS-1 enzyme showed the closest ancestral relationship with OXA-63 from Brachyspira pilosicoli (53% identity) and with putative chromosomal beta-lactamases of Campylobacter spp. (40 to 42% identity). FUS-1 presents all of the conserved structural motifs of class D beta-lactamases. Kinetic analysis revealed that FUS-1 exhibits a narrow substrate profile, efficiently hydrolyzing benzylpenicillin and oxacillin. FUS-1 was poorly inactivated by clavulanate and NaCl. FUS-1 is the first example of a class D beta-lactamase produced by a gram-negative, anaerobic, rod-shaped bacterium to be characterized.

Chromosome- and plasmid-encoded beta-lactamases in Capnocytophaga spp.

Chromosome- and plasmid-encoded CfxA2 and CfxA3 beta-lactamases were detected in Capnocytophaga spp. from oral sources in France, Norway, and the United States. Unidentified chromosome-encoded beta-lactamases were present in Capnocytophaga sputigena. Nucleotide sequence analysis of the CfxA3-encoding plasmid from C. ochracea revealed an unreported insertion sequence (ISCoc1) upstream of the cfxA gene.

Sequence analysis of cfxA2-like beta-lactamases in Prevotella species.

Sixty-two strains of oral (32) and non-oral (30) Prevotella producing beta-lactamases were screened for cfxA by PCR, using an intragenic primer pair. All 62 were cfxA/cfxA2 positive. Fourteen of these strains, representing seven pigmented and seven non-pigmented Prevotella species were submitted to further PCR with specific primers that amplified the whole beta-lactamase structural gene (966 bp). After cloning and sequencing, the deduced amino acid sequences were compared with that of Bacteroides vulgatus CfxA beta-lactamase. All 14 sequences possessed the E272K substitution characteristic of CfxA2. CfxA sensu stricto was not observed in the present series. G83D, F/V189L, W193L and D239Y substitutions were observed more than once, without species specificity. This sequence analysis indicates that most oral and non-oral beta-lactamase-producing Prevotella isolates from French patients produce variants of the CfxA enzyme.