Sequencing and analysis of chromosome 1 of Eimeria tenella reveals a unique segmental organization

Abstract

Eimeria tenella is an intracellular protozoan parasite that infects the intestinal tracts of domestic fowl and causes coccidiosis, a serious and sometimes lethal enteritis. Eimeria falls in the same phylum (Apicomplexa) as several human and animal parasites such as Cryptosporidium, Toxoplasma, and the malaria parasite, Plasmodium. Here we report the sequencing and analysis of the first chromosome of E. tenella, a chromosome believed to carry loci associated with drug resistance and known to differ between virulent and attenuated strains of the parasite. The chromosome--which appears to be representative of the genome--is gene-dense and rich in simple-sequence repeats, many of which appear to give rise to repetitive amino acid tracts in the predicted proteins. Most striking is the segmentation of the chromosome into repeat-rich regions peppered with transposon-like elements and telomere-like repeats, alternating with repeat-free regions. Predicted genes differ in character between the two types of segment, and the repeat-rich regions appear to be associated with strain-to-strain variation.

Figure 1.

E. tenella chromosome 1. (A) The [A+T] content of the sequence (black; 1-kb sliding window) and the ratio of CpG to GpC dinucleotides (red; 1-kb sliding window). (B) The information content (second-order Markov entropy) of the sequence, in a 1-kb sliding window. Values are normalized such that fully repetitive sequence has an information content of 0, and random sequence has an information content of 1. (C) The locations of TGCATGCA motifs (black vertical ticks) or longer [TGCA] tandem repeats (red). (D) Perfect segmental duplications of >50 bp in length along the chromosome (represented by the solid horizontal line); hoops above the line link sequences are duplicated in the same orientation; those below the link are inverted duplications. The color of the hoops indicates the proportion of simple tandem repeats in the duplicated element—(blue) more repetitive, (red) less repetitive; duplicates involving only simple tandem repetitive sequence are not shown. Note that many duplications have only a very short intervening sequence and hence appear as vertical lines. (E) Chromosome 1 with the thickness of the spindle corresponding to the coding density (proportion of nucleotides encoding amino acids; 20-kb center-weighted sliding window; scale at right). Colored bands indicate CAG repeats (red), telomere-like AGGGTTT repeats (green), other simple-sequence repeats (yellow), LINE-related sequences (blue), and gaps (black). The transparent cylinders indicate the feature-poor (P) segments. Physical gaps that are represented as >10 kb in the assembly (and in the corresponding GenBank record) have been condensed to 10 kb in this representation; the distance scale is broken to reflect this. (F) A representative section of a feature-rich (R) segment expanded to show the arrangement of repeat elements (color coding as for panel B) and genes (dark gray; solid segments near the center line indicate complete genes, and narrower wrap-around bands indicate the individual exons). Features are depicted separately for the forward strand (upper) and reverse strand (lower).

Figure 2.

Hexanucleotide content of R- and P-regions. For the R-segment sequence (A) and the P-segment sequence (B), the plots show the relative frequencies (vertical axes) of each possible hexanucleotide. Hexanucleotide sequences are arranged in the horizontal plane, with the homopolymers (T)₆, (A)₆, (C)₆, and (G)₆ at the corners of the plane as indicated. The sequences of the most abundant hexamers in the R-segment sequence (all circular permutations of CAGCAG or its complement) are indicated in A; the peak labeled “Tel” and others of similar height are hexamers from the telomere-like motif AGGGTTT, its complement, and circular permutations thereof.

Figure 3.

Restriction fragment length polymorphisms in P- and R-segments. The figure shows two representative blots. Genomic DNA of E. tenella strains Houghton (H), Weybridge (Wey), or Wisconsin (Wis) was digested with the indicated restriction enzymes, electrophoresed, blotted, and hybridized with radiolabeled probes for P- or R-segment sequences (probe names at top). (A) Shows no size polymorphism between the three strains in the XhoI or HindIII fragments detected by probe Pa; (B) shows a size polymorphism in the HindIII fragment detected by probe Rb.