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Background  During the routine laboratory cultivation of  Lawsonia intracellularis ,  Mycoplasma  contamination has been a frequent problem. When  Mycoplasma  contamination occurs in laboratories that study  L. intracellularis , the cultures must be discarded for 4 reasons: 1)  Mycoplasma  is inevitably concentrated along with  L. intracellularis  during the passage of  L. intracellularis ; 2)  Mycoplasma  inhibits the growth of  L. intracellularis ; and 3) it is impossible to selectively eliminate  Mycoplasma  in  L. intracellularis  cultures. In this study, we observed the contamination of  Mycoplasma  species during  L. intracellularis  cultivation among multiple laboratories.    Results  The presence of a  Mycoplasma  infection in the  L. intracellularis  cultures was verified using polymerase chain reaction (PCR), and a sequence analysis of the partial 16S rRNA and 23S rRNA genes was performed. A PCR-based assay using genus-specific universal primers revealed that 29 (85.3%) of the 34 cultures were contaminated with  Mycoplasma , including 26 with  M. hyorhinis  (89.2%), 2 with  M. orale  (6.9%), and 1 with  M. fermentans  (3.4%). The  Mycoplasma  contamination was not the result of infection with material of pig origin. McCoy cells, which are required for the cultivation of  L. intracellularis , were also ruled out as the source of the  Mycoplasma  contamination.    Conclusions  In this study,  M. hyorhinis  was identified as the most common mollicute that contaminated  L. intracellularis  cultures. Whether  L. intracellularis  enhances the biological properties of  Mycoplasma  to promote infection in McCoy cells is not known. Because the McCoy cell line stocks that were used simultaneously were all negative for  Mycoplasma , and the same worker handled both the McCoy cells to maintain the bacteria and the  L. intracellularis  cultures, it is possible that the  L. intracellularis  cultures are more vulnerable to  Mycoplasma  contamination. Taken together, these results suggest that continuous cultures of  L. intracellularis  must be tested for  Mycoplasma  contamination at regular intervals.  The GenBank accession numbers for the sequences reported in this paper are  JN689375  to  JN689377 .

A multi-laboratory profile of Mycoplasma contamination in Lawsonia intracellularis cultures