Identification of an atypical strain of toxoplasma gondii as the cause of a waterborne outbreak of toxoplasmosis in Santa Isabel do Ivai, Brazil

Abstract

Multilocus DNA sequencing has identified a nonarchetypal strain of Toxoplasma gondii as the causal agent of a waterborne outbreak in Brazil in 2001. The strain, isolated from a water supply epidemiologically linked to the outbreak, was virulent to mice, and it has previously been identified as BrI. Using a serologic assay that detects strain-specific antibodies, we found that 13 (65%) of 20 individuals who were immunoglobulin (Ig) M positive during the outbreak possessed the same serotype as mice infected with the purported epidemic strain. The remaining 7 individuals, plus additional IgM-negative, IgG-positive individuals, possessed 1 of 4 novel serotypes, the most common of which matched the serotype of mice infected with strains isolated from chickens foraging near the outbreak site. The latter strains likely reflect the genetic diversity of T. gondii circulating in highly endemic regions of Brazil. The serotyping assay proved a useful tool for identification of specific individuals infected with the outbreak agent.

Figure 1

Polymorphic sites at the B1 (A), GRA7 (B), and GRA6 (C) loci among archetypal lineages were compared with sequences obtained from the 3 outbreak isolates and 11 strains recovered from chickens grazing in the vicinity of the outbreak in Parana State, Brazil. Consensus sequences were determined by the nucleotide common in at least 2 of the 3 archetypal lineages. Periods (.) denote agreement with the consensus sequence. Dashes (−) denote insertions and deletions (INDELS) in the nucleotide sequence. Nucleotide positions were determined based on the published GenBank sequences for Toxoplasma gondii at the B1 gene (AF179871), GRA7 gene (Y13863.1), and GRA6 gene (AF239283). The first 3 rows contain sequences for archetypal strains I (RH), II (76K), and III (CEP), obtained from laboratory stock; TgCkBr (rows 4–12) designates isolates from chickens in Brazil; outbreak strains (last 3 rows) were obtained from filtered water from a cistern that was epidemiologically linked to the outbreak (strains 1 and 2) and a cat living near the cistern (strain 3). U_I and U_III denote novel alleles that are closely related to types I and III, respectively; u-1, u-2, and u-3 denote novel alleles that are related to, but substantially divergent from, types I, II, and III, respectively.

Figure 2

An enzyme-linked immunosorbent assay plate depicting the 5 distinct serotype patterns identified. The majority of samples exhibited a reactivity pattern consistent with infection by a non–type II strain (labeled “type I/III”), which supported epidemiological evidence. Three additional patterns (labeled “atypical”) were suggestive of infection by nonarchetypal strains, as evidenced by high reactivity to more than 1 allele. In 3 cases, no peptide reactivity was detected, although infection with Toxoplasma gondii was confirmed by SAG1 reactivity (labeled “no reactivity”). Reaction with a type II strain–infected control serum (bottom row) confirmed the specificity of the assay.