A Deep Insight into the Sialome of Rhodnius neglectus, a vector of chagas disease

Background Triatomines are hematophagous insects that act as vectors of Chagas disease. Rhodnius neglectus is one of these kissing bugs found, contributing to the transmission of this American trypanosomiasis. The saliva of hematophagous arthropods contains bioactive molecules responsible for counteracting host haemostatic, inflammatory, and immuneresponses. Methods/Principal Findings Next generation sequencing and mass spectrometry-based protein identification were performed to investigate the content of triatomine R. neglectus saliva.We deposited 4,230 coding DNA sequences (CDS) in GenBank. A set of 636 CDS of proteins of putative secretory nature was extracted from the assembled reads, 73 of them confirmed by proteomic analysis. The sialome of R. neglectus was characterized and serine protease transcripts detected. The presence of ubiquitous protein families was revealed, including lipocalins, serine protease inhibitors, and antigen-5. Metalloproteases, disintegrins, and odorant binding protein families were less abundant. Conclusions/Significance The data presented improve our understanding of hematophagous arthropod sialomes, and aid in understanding hematophagy and the complex interplay among vectors and their vertebrate hosts

Phylogram of lipocalin containing triabin domain from <i>R</i>. <i>neglectus</i> SG transcriptome.

<p>Phylogenetic tree derived from the alignment of <i>R</i>. <i>neglectus</i> CDS and other triatomine lipocalin sequences as described in Methods section. The bar at the bottom represents 20% amino acid substitution. The colored circles indicate each species whose sequences were used: blue, <i>R</i>. <i>neglectus</i> sequences from SG transcriptome; red, <i>R</i>. <i>prolixus</i>; yellow, <i>Triatoma dimidiata</i>; green, <i>Triatoma brasiliensis</i>; dark green, <i>Triatoma matogrossensis</i>; dark blue, <i>T</i>. <i>pallidipennis</i>; purple, <i>Triatoma protacta</i>; magenta, <i>Triatoma infestans</i>; gray, <i>Dipetalogaster maxima</i>.</p

Classification and abundance of proteins from the salivary proteome of <i>R</i>. <i>neglectus</i> based on LC-MS/MS.

<p>Classification and abundance of proteins from the salivary proteome of <i>R</i>. <i>neglectus</i> based on LC-MS/MS.</p

Phylogram of lipocalin containing nitrophorin domain from <i>R</i>. <i>neglectus</i> SG transcriptome.

<p>Phylogenetic tree derived from the alignment of <i>R</i>. <i>neglectus</i> CDS and <i>R</i>. <i>prolixus</i> nitrophorin sequences as described in Methods section. The bar at the bottom represents 20% amino acid substitution. The colored circles indicate each species whose sequences were used: blue, <i>R</i>. <i>neglectus</i> sequences from SG transcriptome and red, <i>R</i>. <i>prolixus</i> sequences from NCBI.</p

Phylogram of Antigen-5 proteins from <i>R</i>. <i>neglectus</i> SG transcriptome.

<p>Phylogenetic tree derived from the alignment of <i>R</i>. <i>neglectus</i> CDS and other insect antigen-5 sequences as described in Methods section. The bar represents 10% amino acid substitution.</p

Kazal-type members from <i>R</i>. <i>neglectus</i> SG transcriptome.

<p>ClustalW alignment of Kazal-type domain-containing members from <i>R</i>. <i>neglectus</i> salivary transcriptome (RN_5563 and RN_549) and other insect kazal-type sequences, identified as described in Methods section. The alignment indicates conserved residues in black and similar residues in gray background, the six conserved cysteines (boxes) and the blue bar indicates the signal peptide indicative of secretion.</p

The ADAMTS sequence from <i>R</i>. <i>neglectus</i> SG transcriptome.

<p>ClustalW alignment of the ADAMTS sequence from <i>R</i>. <i>neglectus</i> salivary transcriptome (RN_11351) and other insect ADAMTS sequences, identified as described in Methods section. The alignment indicates conserved residues in black and similar residues in gray background. The bar indicates the signal peptide indicative of secretion. The boxes limit the family signature sequences showing the determinant residues (black asterisk). The symbols (black hash) above indicate the conserved cysteines.</p

Classification and abundance of coding sequences of putative secretory function extracted from the sialotranscriptome of <i>R</i>. <i>neglectus</i>.

<p>Classification and abundance of coding sequences of putative secretory function extracted from the sialotranscriptome of <i>R</i>. <i>neglectus</i>.</p

Classification and abundance of coding sequences of putative housekeeping function extracted from the sialotranscriptome of <i>R</i>. <i>neglectus</i>.

<p>Classification and abundance of coding sequences of putative housekeeping function extracted from the sialotranscriptome of <i>R</i>. <i>neglectus</i>.</p

The secreted metalloprotease from <i>R</i>. <i>neglectus</i> SG transcriptome.

<p>(A) ClustalW alignment of the secreted metalloprotease from <i>R</i>. <i>neglectus</i> salivary transcriptome (RN_21266) and other metalloproteases sequences, identified as described in Methods section. The alignment indicates conserved residues in black and similar residues in gray background. The blue bar indicates the signal peptide indicative of secretion. The boxes limit the family signature sequences showing the determinant residues (black asterisk). (B) Phylogenetic tree derived from the alignment of <i>R</i>. <i>neglectus</i> CDS and other metalloproteases sequences as described in Methods section. The bar represents 10% amino acid substitution.</p