Fabiola Martino’s scientific contributions

During the period March 2003 – May 2004 at the Laboratory of Clinical Microbiology “Redaelli” LTCRF in Milan, Italy, a total of 529 E. coli, obtained from inpatients of 3 different Long Term Care Rehabilitation Facilities (LTCRFs) in Northern Italy, were processed and 77 ESßLs producers (14.5%) were identified by Vitek System. The results were confirmed by double-disk synergy test with tazobactam (TZP). 61/77 isolates were characterized by higher levels of resistance to cefotaxime (CTX) than to ceftazidime (CAZ). (ß-lactamase production was investigated by analytical isoelectric focusing (IEF) coupled with a bioassay and showed multiple (ß-lactamase bands including one enzyme with pI 8.4 that, in a bioassay, was more active on CTX,ATM than on CAZ. The presence of (ß-lactamase genes was investigated by colony blot hybridization and by PCR amplification of blaTEM, blaSHV and blaCTX-M alleles. 43/61 isolates produced both TEM-1 and CTX-M-type enzymes, 14/61 expressed only CTX-M-type while in 4 cases were found blaCTX-M, blaTEM and blaSHV genes.The remainders (16/77), characterized by high levels of resistance to both CTX and CAZ, produced TEM-1 and SHV-5 enzymes (1/16) and TEM type ESßLs (15/16). Conjugation experiments, performed in liquid medium, confermed that the ESßLs determinants were transferable. Pulsed-field gel electrophoresis profiles of genomic DNA, digested with NotI, were analysed and revealed clonal heterogeneity. Our work confirms the emergence of CTX-M-type enzymes and their spread in Northern Italy also in longterm care and rehabilitation facilities that may be an important reservoir of ES?L producing E. coli.

The VIM- and IMP-type metallo-ß-lactamases (MBLs) are clinically relevant emerging resistance determinants in Gram-negative nosocomial pathogens. To date, MBLs have been reported in acute care hospitals, but their epidemiology in Rehabilitation Facilities (RF) - the largest reservoirs of patients colonized by multidrug-resistant isolates - is largely unknown. This study is aimed to investigate MBL production in Pseudomonas aeruginosa isolates from “Fondazione S. Maugeri” Hospital an RF located in Pavia. 59 non replicate P. aeruginosa strains were isolated between July 2003 - March 2004, from patients incoming from different Italian hospitals. 18/59 resulted resistant to carbapenems. Phenotypic detection of the MBL producers was performed using the E-test and the EPI microdilution method. Molecular methods were used to investigate the MBL determinants and their genetic context.The clonal relatedness of the isolates were studied by PFGE. 12/18 carbapenem-resistant isolates were found to produce a VIM-type MBL. 7/12 were clonally related suggesting a clonal spread, while the 5 others, unrelated to the epidemic profile A, were simultaneously present in the same RF. Preventive and control measures including established microbiological protocols should be applied to detect the MBL producers and to contrast the diffusion of carbapenem-resistant P. aeruginosa in RF.