Q fever, caused by the bacterium C. burnetii, is a zoonosis with a worldwide distribution that affects both humans and animals. From 2007 to 2010, large community outbreaks of Q fever were observed in the Netherlands. In 2008 and 2009 source finding investigations were initiated by several Municipal Health Services, primarily on commercial dairy goat farms, to pinpoint potential sources for Q fever. In that same year, the Food and Consumer Product Safety Authority initiated a project to investigate if petting zoos were a potential source for human Q fever as well. Petting zoos showed insufficient C. burnetii DNA content for molecular typing (qPCR Cq values higher than 33) and were not considered an important source for human Q fever. However, 31 samples from eight out of 57 commercial dairy farms, involved in source finding investigations in 2009, showed a relatively high C. burnetii DNA content based on qPCR data for single copy target com1 (Cq values lower than 33). These samples were selected for molecular typing using a Multi-Locus Variable number of tandem repeat Analysis (MLVA). In this study we show that samples highly positive for C. burnetii DNA can be successfully typed using a multiplex MLVA assay. Three different MLVA types were found, based on six MLVA markers. On seven out of eight locations a single MLVA type was found. On one location a mixture of two types was observed within a number of samples. Our findings show that multiple MLVA types are present in the Netherlands, which is promising for future source finding investigations to identify potential sources. However, only a few different MLVA types have been determined in human and animal samples so far, which makes identifying transmission routes and sources of C. burnetii in the Netherlands still challenging.