Venoarterial extracorporeal membrane oxygenation (VA-ECMO) exposes the patient to infectious complications related to the cannulas or the site of insertion. The aim of the current study was to investigate and compare the prevalence of cannula and membrane oxygenators colonization using three different methods: microbiological culture, scanning electron microscopy, and metagenomic (rRNA 16S analysis). A monocentric prospective study was conducted between December 2017 and June 2018. Consecutive patients undergoing VA-ECMO support for refractory cardiac arrest or cardiogenic shock were included. Ten patients were included with a median age of 64 (52–62) years. Venoarterial extracorporeal membrane oxygenation was inserted for refractory cardiac arrest in five (50%), cardiogenic shock in four (40%), and self-poisoning in one (10%) cases. Microbiological culture of all (8/8, 100%) membrane oxygenators was negative, whereas all (10/10, 100%) were colonized by biofilm, and eight (8/9, 89%) presented bacterial DNA. Three (3/9, 33%) arterial and venous cannulas were positive in culture and seven (7/9, 78%) were colonized by biofilm, respectively. Seven (7/9, 78%) arterial and four (4/9, 44%) venous cannulas presented bacterial DNA. Colonization of cannulas and membranes is more frequent when assessed by electron microscopy or metagenomic analysis than with culture. Membrane oxygenators are more often colonized than cannulas.