Genetic typing of isolates of Rickettsia typhi

Abstract

Murine typhus, which is caused by Rickettsia typhi, has a wide range of clinical manifestations. It has a low mortality rate but may result in meningoencephalitis and interstitial pneumonia in severe cases. Comparisons of complete genome sequences of R. typhi isolates from North Carolina, USA (Wilmington), Myanmar (B9991PP), and Thailand (TH1527) identified only 26 single nucleotide polymorphism (SNP) and 7 insertion-deletion (INDEL) sites in these highly syntenic genomes. Assays were developed to further define the distribution of these variant sites among 15 additional isolates of R. typhi with different histories from Asia, the USA, and Africa. Mismatch amplification mutation assays (MAMA) were validated for 22 SNP sites, while the 7 INDEL sites were analyzed directly on agarose gels. Six SNP types, 9 INDEL types, 11 total types were identified among these 18 isolates. Replicate DNA samples as well as comparisons of isolates with different passage and source histories gave consistent genetic typing profiles. Comparison of the SNP and INDEL markers to R. typhi’s nearest neighbor Rickettsia prowazekii demonstrated that the majority of the SNPs represent intra-species variation that arose post divergence of these two species while several INDEL sites also exhibited intraspecies variability among the R. prowazekii genomes that have been completely sequenced. The assays for the presence of these SNP and INDEL sites, particularly the latter, comprise a low technology gel method for consistently distinguishing R. typhi and R. prowazekii as well as for differentiating genetic types of R. typhi. Author summary: Rickettsia typhi is an obligately intracellular bacterium which is most commonly transmitted by rat fleas to humans and can cause the disease murine (endemic) typhus wherever both humans and infected rats are present. However, this agent is also present in a variety of other arthropod species associated with other vertebrate animals. Its closest relative is Rickettsia prowazekii, the etiologic agent of epidemic typhus (human louse-borne) and sylvatic typhus (associated with flying squirrels and its ectoparasites in the Eastern United States). Both classic louse-borne and flea-borne typhus rickettsiae can cause fatal infections and have the potential for use as biothreat agents as they can be grown in large quantities. We investigated the genetic differences that can be found in different stocks of R. typhi. Our work is important for two reasons. We have developed robust and sensitive methods for distinguishing isolates of R. typhi originating from different continents; these new assays require only simple instrumentation which is available in nearly all biology laboratories. Our results also provided new insights confirming the probable origin of R. typhi in Asia and its likely cosmopolitan spread by rats on ships around the world.

Publication information

Originally published in PLOS Neglected Tropical Diseases on 2022-05-31. Original DOI: 10.1371/journal.pntd.0010354.

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