Implementation of surveillance culture programs and molecular typing are important contributions of Clinical Microbiology to the control of nosocomial infections. This document provides information on collection, transport, preservation, and processing of samples for surveillance culture, as well as the criteria for interpreting and reporting the results of relevant etiologic agents in nosocomial infection. This includes methicillin-resistant Staphylococcus aureus (MRSA), glycopeptide-resistant Enterococcus spp., enterobacteria producing extended-spectrum beta -lactamases (ESBLs), multiresistant Acinetobacter baumannii and carbapenem-resistant Pseudomonas aeruginosa. Details on the available methods for rapid diagnosis are also presented. The information in this document attempts to provide a general approach to the problem and may be considered a starting point for laboratories that are developing their own guidelines, according to needs defined by the multidisciplinary nosocomial infection control team.