(UPDATED July 2021) NUCLEAR 18S rRNA GENE As of July 2021, sequences for 19 isolates of B. mandrillaris have been deposited into the DNA databases (or are available from information in publications or from the authors of reports) for the nuclear 18S rRNA gene. One isolate CDC:V039 has been additionally sequenced independently three times, leading to 23 sequences in the database. Nine isolates have sequences that span almost the complete length of the gene (>1969 bases). Most of the various isolates of B. mandrillaris are very similar to one another in sequence for the 18S rRNA gene, having much less genetic diversity that that seen in Acanthamoeba. As mentioned on the previous page, the lack of variation for the 18S rRNA gene prompted the examination of additional genes to obtain clues to the phylogenetic relationship between isolates. Among the sequences available in the database are 11 sequences obtained as unidentified uncultured eukaryotes in environmental surveys. None of these survey samples contain almost the complete 18S rRNA gene, but the 10 of the 11 exceed 900 basepairs in length. Two of the environmental survey sequences are divergent from clinical samples, showing ~93% sequence identity with clinical isolates of B. mandrillaris. Whether this indicates the existence of a separate taxa closely related to B. mandrillaris (much closer than to either Acanthamoeba or Protacanthameoba) is unclear. Divergence could be due to sequencing errors which sometimes occur in a fraction of environmental uncultured material. MITOCHONDRIAL 16S-LIKE rRNA GENE The sequence of the mitochondrial 16S-like rRNA from B. mandrillaris has proven to be more variable than its nuclear homologue, although levels of variation are still limited. Currently there are 45 sequences representing 38 isolates that have been deposited in the DNA databases for the mitochondrial 16S-like rRNA gene, plus one undeposited sequence. Ten of the sequences come from whole mitochondrial genome sequences, nine of which were released in August 2015. These 10 sequences represent the entire length of the 16S-like rRNA gene sequence and are 1470 and 1487 bp in length (differences represent disagreement of the exact start and end of the rRNA transcript). Other incomplete sequences in the DNA databases range from 1109 bases in length down to 204 nucleotides. In total, 39 of the sequences exceed 800 nucleotides in length. MULTIPLE GENE STUDIES Seven isolates of B. mandrillaris have been studied for both of the rRNA genes. ENVIRONMENTAL ISOLATES Only a single rRNA sequences that clusters with known sequences from Balamuthia (from the nuclear 18S rRNA gene) has been deposited in the DNA databases from uncultured eukaryotes observed during environmental microbiome studies. This is much less than the proportion seen for Vermamoeba, Naegleria, Acanthamoeba or even Protacanthamoeba. This suggests strongly that Balamuthia is truly less frequent in the environment than those other groups of free-living amoebae.