Challenges to get axenic cultures of Trichomonas spp. - A new approach in eradication of contaminants and maintenance of laboratory microbiological cultures.
Contamination of microbiological and cell cultures is a major problem in many scientific and clinical laboratories as well as bioproduct manufacturers worldwide. In the current study we established a rapid (9day) method to detect and eliminate fungal and bacterial contamination in cultures of the unicellular eukaryote Trichomonas spp. The developed method combines identification of the contaminating microorganisms using PCR and sequencing of the 16/18S regions followed by phylogenetic analysis. The next step was a phylogeny-guided selection of antibiotic treatments. We then used a two-step propidium iodide-resorufin assay to test the effect of selected antibiotics. The result was a quick and worthwhile purification of trichomonad laboratory cultures. Our workflow may also be implemented to obtain new isolates of trichomonads from clinical samples if initial broad-spectrum antibiotic therapy fails.
Division of Pharmacognosy, Department of Medicinal Chemistry, Uppsala University, Uppsala 75123, Sweden; Volga State University of Technology, Yoshkar-Ola 424000, Russia. Electronic address: anna.koptina@fkog.uu.se.
Anna
A
Koptina
Division of Pharmacognosy, Department of Medicinal Chemistry, Uppsala University, Uppsala 75123, Sweden.
Åke
Å
Strese
Division of Pharmacognosy, Department of Medicinal Chemistry, Uppsala University, Uppsala 75123, Sweden.
Anders
A
Backlund
Division of Pharmacognosy, Department of Medicinal Chemistry, Uppsala University, Uppsala 75123, Sweden; Department of Virology, Immunobiology, and Parasitology, National Veterinary Institute (SVA), Uppsala 75651, Sweden.
Cecilia
C
Alsmark
D000890:Anti-Infective Agents / Q000494:pharmacology; D061245:Axenic Culture; D001419:Bacteria / Q000145:classification / Q000187:drug effects / Q000302:isolation & purification; D004275:DNA, Ribosomal / Q000737:chemistry / Q000235:genetics; D005658:Fungi / Q000145:classification / Q000187:drug effects / Q000302:isolation & purification; D010274:Parasitology / Q000379:methods*; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012337:RNA, Ribosomal, 18S / Q000235:genetics; D017422:Sequence Analysis, DNA; D013997:Time Factors; D014244:Trichomonas / Q000254:growth & development* / Q000302:isolation & purification*
D000890:Anti-Infective Agents; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S; D012337:RNA, Ribosomal, 18S
Antibiotic; Contamination; Phylogeny; Propidium iodide; Resorufin; Trichomonas spp.