Acquisition of exogenous haem is essential for tick reproduction
- Biology Centre of the Czech Academy of Sciences, Czech Republic
- University of South Bohemia, Czech Republic
- Czech Academy of Sciences, Czech Republic
- Universidade Federal do Rio de Janeiro, Brazil
- Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular, Brazil
Figures
Figure 1 with 5 supplements
Evolution of haem biosynthetic and degradative pathways.
(A) General scheme of haem biosynthetic and degradative pathways in the eukaryotic cell. Haem biosynthesis (upper) is a series of eight reactions beginning in the mitochondria by condensation of …
Figure 1—figure supplement 1
Phylogenetic tree of selected coproporphyrinogen-III oxidases.
Unrooted tree of coproporphyrinogen-III oxidase (CPOX) amino acid sequences reconstructed using the Neighbor Joining method (NJ) based on alignment using ClustalX. The Ixodes scapularis and Ixodes …
Figure 1—figure supplement 2
Phylogenetic tree of selected protoporphyrinogen oxidases.
Unrooted tree of protoporphyrinogen oxidase (PPOX) amino acid sequences reconstructed using the Neighbor Joining method (NJ) based on alignment using ClustalX. The Ixodes scapularis, Ixodes ricinus, …
Figure 1—figure supplement 3
Phylogenetic tree of selected ferrochelatases.
Unrooted tree of ferrochelatase (FECH) amino acid sequences reconstructed using the Neighbor Joining method (NJ) based on alignment using ClustalX. The Ixodes scapularis and Ixodes ricinus FECHs …
Figure 1—figure supplement 4
Phylogenetic tree of selected 5-aminolevulinate synthases.
Unrooted tree of 5-aminolevulinate synthase (ALAS) amino acid sequences reconstructed using the Neighbor Joining method (NJ) based on alignment using ClustalX. The ISCW020754 annotated in the Ixodes …
Figure 1—figure supplement 5
Phylogenetic tree of selected uroporphyrinogen decarboxylases.
Unrooted tree of uroporphyrinogen decarboxylase (UROD) amino acid sequences reconstructed using the Neighbor Joining method (NJ) based on alignment using ClustalX. The ISCW020804 annotated in the Ixo…
Figure 2 with 2 supplements
Impact of dietary haemoglobin on tick feeding, oviposition, embryogenesis, and larval hatching.
(Membrane feeding) - membrane feeding in vitro of Ixodes ricinus females on whole blood (Blood-fed), serum (Serum-fed) and on serum supplemented with 10% bovine haemoglobin (Serum + 10% Hb). For …
Figure 2—figure supplement 1
Diets used for tick membrane feeding and faecal examination.
(A) Females of I. ricinus were membrane fed until full engorgement (FE) using whole blood, serum, and serum supplemented with 10% (physiological concentration) of pure bovine haemoglobin (Serum+Hb). …
Figure 2—figure supplement 2
Rescue experiments with sub-physiological levels of haemoglobin.
Embryonal development and larval hatching was fully rescued in I. ricinus females fed on serum supplemented with 1% or 0.1% bovine haemoglobin. (Embryogenesis) – microscopic examination of embryonal …
Figure 3 with 3 supplements
Determination of haemoglobin-derived nutrients in ticks (haem, amino acids, iron).
(A) Levels of haem b were determined by HPLC in egg homogenates from ticks fed on whole blood (BF) serum (SF), and serum supplemented with 10%, 1% or 0.1% bovine haemoglobin (S+10%Hb, S+1%Hb and …
Figure 3—figure supplement 1
HPLC analysis of haem b in tick egg homogenates.
Homogenates prepared from 10 mg of eggs were collected from three independent egg clutches laid by I. ricinus females fed on different diets. Representative chromatograms are shown detecting haem b …
Figure 3—figure supplement 2
Full appearance of SDS-PAGE and Western blot analyses shown in the Figure 3.
SDS-PAGE analyses were carried out on homogenates prepared from three independent tissue preparations from ticks fed on whole blood (BF) or serum (SF). Protein profiles were visualized using the TGX …
Figure 3—figure supplement 3
Detection of biliverdin IX derivatives in Ixodes ricinus and Aedes aegypti.
The HPLC using a diode array detector was set to enable a simultaneous determination of haem b and biliverdin IX compounds at wavelengths of 375 nm and 660 nm, respectively. For details, see …
Figure 4
Appearance of the tick gut, digest cells, and ovaries from blood- and serum-fed ticks.
Whole guts from blood-fed (BF) and serum-fed (SF) partially engorged females (fed for 6 days) were dissected and semi-thin sections of digest cells were prepared and stained with toluidine blue. L - …
Figure 5 with 1 supplement
IrCP3 is the major haem-binding protein in I. ricinus haemolymph.
(A-C) Ir-CP3 and haem levels in haemolymph collected from blood-fed (BF) and serum-fed (SF) partially engorged females. (A) Absorbance spectra of haemolymph samples from BF and SF females. (B) …
Figure 5—figure supplement 1
Stage and tissue expression of I. ricinus haemolymph carrier protein (IrCP3).
(A) qPCR analyses of ir-cp3 expression in developmental stages of I. ricinus. (B) qPCR analyses of ir-cp3 expression in tissues dissected from fully engorged females. Data were obtained from three …
Figure 6 with 3 supplements
Vitellins are the major haem-binding proteins in tick ovaries.
(A-B) Haem accumulation in tick ovaries occur concurrently with the appearance of vitellins. Ovaries were dissected from I. ricinus females at subsequent time-points after detachment (AD) from the …
Figure 6—figure supplement 1
SDS-PAGE and Western blot analyses of ovary homogenates from I. ricinus.
Ovaries were dissected from I. ricinus females at subsequent time-points after detachment (AD) from the host: FE - fully-engorged; 3 AD, 6 AD, 8 AD - 3, 6, and 8 days AD, respectively. Protein …
Figure 6—figure supplement 2
Stage and tissue expression of I. ricinus vitellogenin 1 (IrVg1) and vitellogenin 2 (IrVg2).
Stage and tissue expression of I. ricinus vitellogenin 1 (IrVg1) and vitellogenin 2 (IrVg2). (A) qPCR analyses of ir-vg1 and ir-vg2 expression in developmental stages of I. ricinus. (B) qPCR …
Figure 6—figure supplement 3
RNAi-mediated silencing of I. ricinus vitellogenin 1 and 2.
Unfed I. ricinus females were pre-injected with gfp dsRNA (gfp), ir-vg1 dsRNA (ir-vg1 KD), ir-vg2 dsRNA (ir-vg2 KD), and ir-cp3 dsRNA (ir-cp3 KD), allowed to feed naturally on guinea pigs and …
Author response image 1
Table 1: Overview of the feeding and egg laying parameters of membrane-fed I. ricinus females.
https://doi.org/10.7554/eLife.12318.026
Author response image 2
Experimental feeding in vitro of I. ricinus females on serum supplemented with myoglobin or haemin.
Ticks were membrane fed in vitro on bovine serum (S). Pure equine myoglobin (Sigma, M0630) or haemin (Sigma, H9039) of specified concentrations were added to the serum diet from the 6th day of …
Author response image 3
Expression of coproporphyrinogen-III oxidase (ir-cpox), protoporphyrinogen oxidase (ir-ppox), and ferrochelatase (ir-fech) in the tick Ixodes ricinus.
(A) Expression profiles of nymphal and female adult I. ricinus stages over initial phases of feeding from available RNAseq data (Kotsyfakis et al., 2015). SG – salivary glands; MG – gut; N – nymph; …
Author response image 4
Increased transferrin levels in serum led to higher levels of ferritin 1 in tick gut.
Ticks were fed on serum or on serum supplemented with 3 mg/ml of bovine holo-Transferrin (Sigma, T1283). This addition increased the concentration of transferrin in serum approximately 2-fold, …
Additional files
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Supplementary file 1
Prediction of genes coding for haemoproteins in the genome of I. scapularis and their putative function in tick metabolism.
- https://doi.org/10.7554/eLife.12318.023
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Supplementary file 2
Design, sequences, and sequence similarities of recombinant Vitellogenin_N domains of IrCP3, IrVg1, and IrVg2 used for raising specific antibodies.
- https://doi.org/10.7554/eLife.12318.024
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Supplementary file 3
Oligonucleotides used in this work.
- https://doi.org/10.7554/eLife.12318.025
