Bacteriolytic activity in saliva of the hematophagous Triatoma infestans (Reduviidae) and novel characterization and expression site of a third lysozyme

• Published in: Archives of insect biochemistry and physiology Vol. 113; no. 3; pp. e22013 - n/a
• Main Authors: Meiser, Christian K., Klenner, Lars, Balczun, Carsten, Schaub, Günter A.
• Format: Journal Article
• Language: English
• Published: United States Wiley Subscription Services, Inc 01-07-2023
• Subjects: antibacterial compounds; Exocrine glands; Feeding; Gene expression; Insects; Instars; Lysis; Lysozyme; lysozyme genes; Micrococcus luteus; Oligonucleotides; pH effects; Polymerase chain reaction; Reverse transcription; Saliva; Salivary gland; Salivary glands; Triatoma infestans; zymogram; saliva; Triatoma infestans; antibacterial compounds; zymogram; lysozyme genes
• ISSN: 0739-4462; 1520-6327
• DOI: 10.1002/arch.22013

Saliva of hematophagous insects contains many different compounds, mainly acting as anticoagulants. Investigating the bacteriolytic compounds of the saliva of the bloodsucking Triatoma infestans photometrically between pH 3 and pH 10 using unfed fifth instars and nymphs up to 15 days after feeding, we found bacteriolytic activity against lyophilized Micrococcus luteus was stronger at pH 4 and pH 6. After feeding, the activity level at pH 4 was unchanged, but at pH 6 more than doubled between 3 and 7 days after feeding. In zymographs of the saliva and after incubation at pH 4, bacteriolytic activity against Micrococcus luteus was present at eight lysis zones between 14.1 and 38.5 kDa, showing the strongest activity at 24.5 kDa. After incubation at pH 6, lysis zones only appeared at 15.3, 17, and 31.4 kDa. Comparing zymographs of the saliva of unfed and fed nymphs, bacteriolytic activity at 17 kDa increased after feeding. In total nine lysis bands appeared, also at >30 kDa, so far unreported in the saliva of triatomines. Reverse transcription polymerase chain reaction using oligonucleotides based on the previously described lysozyme gene of T. infestans, TiLys1, verified expression of genes encoding TiLys1 and TiLys2 in the salivary glands, but also of an undescribed third lysozyme, TiLys3, of which the cloned cDNA shares characteristics with other c‐type lysozymes of insects. While TiLys1 was expressed in the tissue of all three salivary glands, transcripts of TiLys2 and of TiLys3 seem to be present only in the gland G1 and G3, respectively.
Highlights
In zymographs of the saliva of Triatoma infestans, in total nine lysis zones of bacteria were present at 14–39 kDa, and bacteriolytic compounds at >30 kDa are unknown for triatomines.
Polymerase chain reaction verified an expression of a so far undescribed third lysozyme, named TiLys3.
Bacteriolytic activity after zymography of saliva obtained from unfed nymphs or 7 days after feeding and after an incubation at pH 4 and pH 6, followed by staining and additional incubation in water. Molecular masses of marker proteins are displayed on the left. Asterisks indicate lysis zones present after the respective incubation