A comparison of two membrane-systems for the in vitro blood-feeding ofSimulium damnosumTheobald s. 1. (Dipt., Simuliidae)

Blood feeding insects have to rely on the protein content of mammalian blood to insure reproduction. A substantial quantity of protein is provided by hemoglobin present in erythrocytes. Access to hemoglobin is accomplished only via erythrocyte lysis. It has been shown that midgut homogenates from the blood feeding stable fly, Stomoxys calcitrans, contain free fatty acids and it was proposed that these detergent-like compounds play a major role as hemolysins in the digestive physiology of this species. More recently sphingomyelinase activity was detected in midgut preparations of this fly, which would provide a potential tool for the enzymatic cleavage of the erythrocyte's membrane sphingomyelin. The action of specific hemolytic factors should affect the erythrocyte's morphology. The shape of bovine erythrocytes undergoing in vitro hemolysis by crude midgut homogenates from the stable fly was examined by scanning and transmission electron microscopy.

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Biological and physiological characterization of in vitro blood feeding in nymph and adult stages of Ornithodoros turicata (Acari: Argasidae)

Journal of Insect Physiology ◽

10.1016/j.jinsphys.2015.03.005 ◽

2015 ◽

Vol 75 ◽

pp. 73-79 ◽

Cited By ~ 11

Author(s):

Hongyuan Zheng ◽

Andrew Y. Li ◽

Pete D. Teel ◽

Adalberto A. Pérez de León ◽

Janakiram Seshu ◽

...

Keyword(s):

Blood Feeding ◽

Physiological Characterization

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Biomimetic Model Membrane Systems Serve as Increasingly Valuable in Vitro Tools

Advances in Biomimetics ◽

10.5772/13932 ◽

2011 ◽

Author(s):

Mary T. ◽

Jennifer K. ◽

Elmar J.

Keyword(s):

Model Membrane ◽

Membrane Systems

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Blood Digestion by Trypsin-Like Serine Proteases in the Replete Lyme Disease Vector Tick, Ixodes scapularis

Insects ◽

10.3390/insects11030201 ◽

2020 ◽

Vol 11 (3) ◽

pp. 201

Author(s):

Jeremiah Reyes ◽

Cuauhtemoc Ayala-Chavez ◽

Arvind Sharma ◽

Michael Pham ◽

Andrew B. Nuss ◽

...

Keyword(s):

Lyme Disease ◽

Blood Meal ◽

Serine Proteases ◽

Ixodes Scapularis ◽

Life Stage ◽

Blood Feeding ◽

Eastern United States ◽

Blood Digestion ◽

Hemoglobin Degradation

Ixodes scapularis is the major vector of Lyme disease in the Eastern United States. Each active life stage (larva, nymph, and adult) takes a blood meal either for developing and molting to the next stage (larvae and nymphs) or for oviposition (adult females). This protein-rich blood meal is the only food taken by Ixodes ticks and therefore efficient blood digestion is critical for survival. Studies in partially engorged ticks have shown that the initial stages of digestion are carried out by cathepsin proteases within acidic digestive cells. In this study, we investigated the potential role of serine proteases in blood digestion in replete ticks. RNA interference was used for functional analysis and a trypsin-benzoyl-D, L-arginine 4-nitoanilide assay was used to measure active trypsin levels. Hemoglobinolytic activity was determined in vitro, with or without a serine protease inhibitor. Our data suggest that trypsin levels increase significantly after repletion. Knockdown of serine proteases negatively impacted blood feeding, survival, fecundity, levels of active trypsin in the midgut, and resulted in lower hemoglobin degradation. Incubation of midgut extract with a trypsin inhibitor resulted in 65% lower hemoglobin degradation. We provide evidence of the serine proteases as digestive enzymes in fully engorged, replete females. Understanding the digestive profile of trypsin during blood meal digestion in I. scapularis improves our understanding of the basic biology of ticks and may lead to new methods for tick control.

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Bioinspired membrane-based systems for a physical approach of cell organization and dynamics: usefulness and limitations

Interface Focus ◽

10.1098/rsfs.2015.0038 ◽

2015 ◽

Vol 5 (4) ◽

pp. 20150038 ◽

Cited By ~ 35

Author(s):

Thibaut J. Lagny ◽

Patricia Bassereau

Keyword(s):

Cell Membranes ◽

Cell Communication ◽

Basic Research ◽

Model Membrane ◽

Membrane Systems ◽

Vital Functions ◽

Cell Components ◽

Potential Applications ◽

Long Time

Being at the periphery of each cell compartment and enclosing the entire cell while interacting with a large part of cell components, cell membranes participate in most of the cell's vital functions. Biologists have worked for a long time on deciphering how membranes are organized, how they contribute to trafficking, motility, cytokinesis, cell–cell communication, information transport, etc., using top-down approaches and always more advanced techniques. In contrast, physicists have developed bottom-up approaches and minimal model membrane systems of growing complexity in order to build up general models that explain how cell membranes work and how they interact with proteins, e.g. the cytoskeleton. We review the different model membrane systems that are currently available, and how they can help deciphering cell functioning, but also list their limitations. Model membrane systems are also used in synthetic biology and can have potential applications beyond basic research. We discuss the possible synergy between the development of complex in vitro membrane systems in a biological context and for technological applications. Questions that could also be discussed are: what can we still do with synthetic systems, where do we stop building up and which are the alternative solutions?

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The MF6p/FhHDM-1 Major Antigen Secreted by the Trematode Parasite Fasciola hepatica Is a Heme-binding Protein

Journal of Biological Chemistry ◽

10.1074/jbc.m113.499517 ◽

2013 ◽

Vol 289 (3) ◽

pp. 1441-1456 ◽

Cited By ~ 21

Author(s):

Victoria Martínez-Sernández ◽

Mercedes Mezo ◽

Marta González-Warleta ◽

María J. Perteguer ◽

Laura Muiño ◽

...

Keyword(s):

Fasciola Hepatica ◽

Binding Protein ◽

Blood Feeding ◽

Reversed Phase ◽

Heme Binding ◽

Free Heme ◽

Major Antigen ◽

Parenchymal Cells ◽

Heme Binding Protein

Blood-feeding parasites have developed biochemical mechanisms to control heme intake and detoxification. Here we show that a major antigen secreted by Fasciola hepatica, previously reported as MF6p, of unknown function (gb|CCA61804.1), and as FhHDM-1, considered to be a helminth defense molecule belonging to the family of cathelicidin-like proteins (gb|ADZ24001.1), is in fact a heme-binding protein. The heme-binding nature of the MF6p/FhHDM-1 protein was revealed in two independent experiments: (i) immunopurification of the secreted protein·heme complexes with mAb MF6 and subsequent analysis by C8 reversed-phase HPLC and MS/MS spectrometry and (ii) analysis of the binding ability of the synthetic protein to hemin in vitro. By immunohistochemistry analysis, we have observed that MF6p/FhHDM-1 is produced by parenchymal cells and transported to other tissues (e.g. vitellaria and testis). Interestingly, MF6p/FhHDM-1 is absent both in the intestinal cells and in the lumen of cecum, but it can be released through the tegumental surface to the external medium, where it binds to free heme molecules regurgitated by the parasite after hemoglobin digestion. Proteins that are close analogs of the Fasciola MF6p/FhHDM-1 are present in other trematodes, including Clonorchis, Opistorchis, Paragonimus, Schistosoma, and Dicrocoelium. Using UV-visible spectroscopy and immunoprecipitation techniques, we observed that synthetic MF6p/FhHDM-1 binds to hemin with 1:1 stoichiometry and an apparent Kd of 1.14 × 10−6m−1. We also demonstrated that formation of synthetic MF6p/FhHDM-1·hemin complexes inhibited hemin degradation by hydrogen peroxide and hemin peroxidase-like activity in vitro. Our results suggest that MF6p/FhHDM-1 may be involved in heme homeostasis in trematodes.

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Effects of Formic Acid on Amblyomma americanum (Ixodida: Ixodidae) Larvae and Nymphs

Journal of Medical Entomology ◽

10.1093/jme/tjaa037 ◽

2020 ◽

Vol 57 (4) ◽

pp. 1184-1192

Author(s):

Allan T Showler ◽

Bailee N Dorsey ◽

Ryan M Caesar

Keyword(s):

Formic Acid ◽

Varroa Destructor ◽

Direct Contact ◽

Acid Treatment ◽

Blood Feeding ◽

Amblyomma Americanum ◽

Alternative Control ◽

Temporary Immersion ◽

Varroa Mites

Abstract Ixodids are blood-feeding ectoparasitic vectors of many disease agents that infect humans, livestock, and wild animals. As ixodid resistance to conventional synthetic acaricides becomes increasingly problematic, natural products are receiving greater attention as possible alternative control tactics. Formic acid, produced by ants, is a commercially available product for fumigating varroa mites, Varroa destructor Anderson & Trueman, infesting honey bee, Apis mellifera L., hives, and it has been reported to repel ixodids. Lone star tick, Amblyomma americanum (L.), larvae and nymphs were used as a model ixodid to investigate deterrent, repellent, and lethal effects of formic acid as a fumigant and contact toxin in vitro in the laboratory. Although formic acid failed to deter or repel A. americanum, it was highly toxic as a fumigant to larvae at a 1% concentration even when exposure was limited to 5 min. Contact by crawling on wet, moist, and dry treated substrates under ventilated conditions causes >90% mortality to larvae in 5% formic acid concentration treatments within 30–120 min, and temporary immersion killed ≈60% of the larvae by 24 h after they were removed from the 5% formic acid treatment solution. Substantial nymphal mortality occurred after 1–1.5 h following exposure to substrate treated with the 10% concentration and immersion killed ≈45% of the nymphs. It appears that formic acid volatiles are more lethal to A. americanum immatures than direct contact with the external integument.

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In Silico Characterization and Structural Modeling of Dermacentor andersoni p36 Immunosuppressive Protein

Advances in Bioinformatics ◽

10.1155/2018/7963401 ◽

2018 ◽

Vol 2018 ◽

pp. 1-12 ◽

Cited By ~ 2

Author(s):

Martin Omulindi Oyugi ◽

Johnson Kangethe Kinyua ◽

Esther Nkirote Magiri ◽

Milcah Wagio Kigoni ◽

Evenilton Pessoa Costa ◽

...

Keyword(s):

Vaccine Development ◽

Dynamic Properties ◽

3D Structure ◽

Blood Feeding ◽

Rhipicephalus Appendiculatus ◽

Economic Losses ◽

Dermacentor Andersoni ◽

Laboratory Confirmation ◽

Tick Vaccine

Ticks cause approximately $17–19 billion economic losses to the livestock industry globally. Development of recombinant antitick vaccine is greatly hindered by insufficient knowledge and understanding of proteins expressed by ticks. Ticks secrete immunosuppressant proteins that modulate the host’s immune system during blood feeding; these molecules could be a target for antivector vaccine development. Recombinant p36, a 36 kDa immunosuppressor from the saliva of female Dermacentor andersoni, suppresses T-lymphocytes proliferation in vitro. To identify potential unique structural and dynamic properties responsible for the immunosuppressive function of p36 proteins, this study utilized bioinformatic tool to characterize and model structure of D. andersoni p36 protein. Evaluation of p36 protein family as suitable vaccine antigens predicted a p36 homolog in Rhipicephalus appendiculatus, the tick vector of East Coast fever, with an antigenicity score of 0.7701 that compares well with that of Bm86 (0.7681), the protein antigen that constitute commercial tick vaccine Tickgard™. Ab initio modeling of the D. andersoni p36 protein yielded a 3D structure that predicted conserved antigenic region, which has potential of binding immunomodulating ligands including glycerol and lactose, found located within exposed loop, suggesting a likely role in immunosuppressive function of tick p36 proteins. Laboratory confirmation of these preliminary results is necessary in future studies.

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The early gene Broad is involved in the ecdysteroid hierarchy governing vitellogenesis of the mosquito Aedes aegypti

Journal of Molecular Endocrinology ◽

10.1677/jme.1.01531 ◽

2004 ◽

Vol 33 (3) ◽

pp. 743-761 ◽

Cited By ~ 42

Author(s):

L Chen ◽

J Zhu ◽

G Sun ◽

A S Raikhel

Keyword(s):

Aedes Aegypti ◽

Fat Body ◽

Regulatory Region ◽

Blood Feeding ◽

Early Gene ◽

Gene Encoding ◽

Obvious Effect ◽

Early Genes ◽

Old Females

The broad (br ) gene, encoding a family of C2H2 type zinc-finger DNA-binding proteins, has been shown to act as a crucial member of the 20-hydroxyecdysone (20E) regulatory hierarchy in the fruitfly, Drosophila melanogaster and the moth, Manduca sexta. In this study, we have shown that the br gene is involved in the 20E-regulatory hierarchy controlling vitellogenesis in the mosquito, Aedes aegypti. Unlike E74 and E75 early genes, expression of br was activated in previtellogenic females, during a juvenile hormone (JH)-dependent period. The levels of Z1, Z2 and Z4 isoform mRNA were elevated in the fat body of 2-day-old females after in vitro exposure to JH III. However, JH III repressed 20E activation of br in 3-to 5-day-old females, indicating a switch in hormonal commitment. Expression of Z1, Z2 and Z4 was stimulated after blood feeding in both vitellogenic tissues, the fat body and the ovary, corresponding to peaks of ecdysteroid titers. In the fat body, the mRNA profiles of these three isoforms correlated well with those of yolk protein precursor (YPP) genes. These BR isoforms were activated by 20E in fat bodies cultured in vitro and behaved as early genes, with a self-repressive autoregulatory loop that can be blocked by the protein inhibitor, cyclohexamide. Multiple binding sites for all four BR isoforms were present in the 5′-regulatory region of the major YPP gene, vitellogenin (Vg). Effects of BR isoforms on the expression of Vg have been demonstrated by cell transfection analysis. In particular, BR isoforms by themselves had no effects on the Vg promoter. However, Z1 and Z4 each repressed Aedes aegypti ecdysone receptor (EcR)/Ultraspiracle (USP)-mediated 20E activation of the Vg promoter, while Z2 enhanced activation of the Vg promoter by AaEcR/AaUSP in the presence of 20E. Z3 had no obvious effect in the same experiment. These results suggested that BR isoforms are essential for proper activation and termination of the Vg gene in response to 20E. Overall, our study implicated br in the regulation of mosquito vitellogenesis.

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Blood Digestion by Trypsin-Like Serine Protease in the Replete Lyme Disease Vector Tick, Ixodes scapularis

10.20944/preprints201910.0026.v2 ◽

2020 ◽

Author(s):

Jeremiah Reyes ◽

Cuauhtemoc Ayala-Chavez ◽

Michael Pham ◽

Arvind Sharma ◽

Andrew Nuss ◽

...

Keyword(s):

Lyme Disease ◽

Serine Protease ◽

Blood Meal ◽

Serine Proteases ◽

Ixodes Scapularis ◽

Life Stage ◽

Blood Feeding ◽

Blood Digestion ◽

Hemoglobin Degradation

Ixodes scapularis is the major vector of Lyme disease in the eastern United States. Each active life stage (larva, nymph, and adult) takes a blood meal either for developing and molting to the next stage (larvae and nymphs) or for oviposition (adult females). This protein-rich blood meal is the only food taken by Ixodes ticks and therefore blood digestion is very important for tick survival. Most studies on blood digestion in ticks have shown that the initial stages of digestion are carried out by cathepsin proteases within acidic digestive cells. However, most of these studies have focused on partially engorged ticks. In other hematophagous arthropods, the serine proteases play an important role in blood protein degradation. In this study, we determined transcript expression of four I. scapularis serine proteases with previously characterized roles in blood digestion. RNA interference was used for functional analysis and a trypsin-benzoyl-D, L-arginine 4-nitoanilide assay was used to measure active trypsin levels. An in vitro hemoglobinolytic assay was performed with or without serine protease inhibitor. Our data suggest that trypsin levels increase significantly after blood feeding and peaked in larvae, nymphs, and adults at 3, 1, and 1 day post host detachment, respectively. The knockdown of three previously identified serine proteases by RNAi negatively impacted blood intake, survival, fecundity, levels of active trypsin in the gut and resulted in lower hemoglobin degradation in vitro. A trypsin inhibitor, PMSF, blocked the action of trypsin in the gut extract resulting in 65% lower hemoglobin degradation. We provide evidence of the serine proteases as digestive enzymes in fully engorged, replete females. Our data also demonstrated that in addition to blood digestion, these serine proteases might have a role in blood feeding success in I. scapularis.

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