Purification and cloning of the salivary nitrophorin from the hemipteran Cimex lectularius.

1998 ◽  
Vol 201 (18) ◽  
pp. 2659-2664 ◽  
Author(s):  
J G Valenzuela ◽  
J M Ribeiro
Keyword(s):  
Molecular Mass ◽  
High Performance ◽  
Blood Feeding ◽  
Protein Sequencing ◽  
Cimex Lectularius ◽  
Base Pairs ◽  
Clone Sequence ◽  
Internal Peptide ◽  
Vertebrate Hosts ◽  
Inositol Phosphatases

Cimex lectularius and Rhodnius prolixus contain salivary nitric oxide (NO) that may help them to feed on their vertebrate hosts by promoting vasodilation and inhibiting platelet aggregation. Salivary NO is associated with heme proteins (nitrophorins) that store and transport NO from the insect salivary glands to the skin of the host. In this study, the salivary nitrophorin of Cimex lectularius was purified by DEAE chromatography and reverse-phase high-performance liquid chromatography. The purified nitrophorin had a molecular mass of 32.9 kDa. The DEAE-purified hemoprotein was able to bind NO, and this binding shifted the absorption maximum from 388 nm to 438 nm. The ratio of heme to apoprotein was estimated to be of 1:1. A cDNA clone of 1079 base pairs was sequenced and was found to code for a protein with a molecular mass of 31.7 kDa. The clone sequence was in agreement with the internal peptide sequences obtained from the purified protein. Sequencing of the isolated clone indicates high similarity to several inositol phosphatases; however, no significant similarities emerged when the sequence of C. lectularius nitrophorin was compared with that of R. prolixus nitrophorin, the only other nitrophorin known in insect saliva. Because C. lectularius and R. prolixus belong to two different families of Hemiptera that evolved independently to blood feeding, a case is made for the convergent evolution of these two insect nitrophorins.

Identification of Bloodmeals from Sand Flies (Diptera: Psychodidae) Collected in the Parque Nacional do Viruá, State of Roraima, Brazil

2021 ◽  
Author(s):  
Bruno Leite Rodrigues ◽  
Glaucilene da Silva Costa ◽  
Paloma Helena Fernandes Shimabukuro
Keyword(s):  
Blood Feeding ◽  
Sand Flies ◽  
Feeding Preferences ◽  
Light Traps ◽  
Dasypus Novemcinctus ◽  
Search Tool ◽  
Vertebrate Hosts ◽  
Host Blood ◽  
Clustering Pattern ◽  
Species Being

Abstract The transmission of pathogens that cause leishmaniases occurs by the bite of female sand flies (Diptera: Psychodidae) in their vertebrate hosts, which makes the identification of their bloodmeal sources an important step for the control and epidemiology of these diseases. In Brazil, the state of Roraima has a great diversity of sand flies, vertebrate hosts, and protozoan Leishmania, but little is known about the host blood-feeding preferences of sand flies. Thus, we evaluated the bloodmeal sources of sand flies collected from their sylvatic habitats in Parque Nacional do Viruá, Roraima. Fieldwork was carried-out between 13th and 18th August 2019 using CDC light traps. Sand flies were slide-mounted and morphologically identified using the head and last segments of the abdomen. Engorged females had their DNA extracted, followed by amplification and sequencing of the cytochrome b (cytb) molecular marker for vertebrates. Sequences were analyzed and compared with those from GenBank using the BLASTn search tool, in addition to the reconstruction of a phylogenetic tree to demonstrate the clustering pattern of these sequences. A total of 1,209 sand flies were identified, comprising 20 species, in which the most abundant were Psychodopygus ayrozai (Barretto and Coutinho) (42.10%) and Psychodopygus chagasi (Costa Lima) (26.22%). Bloodmeal source identification was successfully performed for 34 sand flies, that confirm four vertebrate species, being the most abundant the armadillo Dasypus novemcinctus Linnaeus, 1758 (Cingulata: Dasypodidae).

The Behavioral Response to Heat in the Common Bed Bug,Cimex lectularius (Hemiptera: Cimicidae)

2021 ◽  
Author(s):  
Raymond Berry
Keyword(s):  
Petri Dish ◽  
Cimex Lectularius ◽  
Bed Bugs ◽  
Bed Bug ◽  
Host Interaction ◽  
Video Images ◽  
Spherical Ball ◽  
The Common ◽  
Vertebrate Hosts

AbstractThe bed bug, Cimex lectularius L., is a common ectoparasite found to live among its vertebrate hosts. Antennal segments in bugs are critical for sensing multiple cues in the environment for survival. To determine whether the thermo receptors of bed bugs are located on their antennae; innovative bioassays were created to observe the choice between heated and unheated stimuli and to characterize the response of bugs to a heat source. Additionally, the effect of complete antenectomized segments on heat detection were evaluated. Heat, carbon dioxide, and moisture are cues that are found to activate bed bug behavior; a temperature at 38°C was used to assess the direction/degree at which the insect reacts to the change in distance from said stimulus. Using a lightweight spherical ball suspended by air through a vacuum tube, bed bugs and other insects are able to move in 360° while on a stationary point. Noldus EthoVision XT was used to capture video images and to track the bed bugs during 5-min bioassays. A bioassay was created using four Petri dish arenas to observe bed bug attraction to heat based on antennae segments at 40°C. The purpose of this study was to evaluate the effects of heat on complete antenectomized segments of the antennae. The results in this experiment suggest that bed bugs detect and are attracted to heat modulated by nutritional status. Learning the involvement of antennae segments in heat detection will help identify the location and role of thermoreceptors for bed bug host interaction.

scholarly journals Cadmium-Responsive Thiols in the Ectomycorrhizal Fungus Paxillus involutus

2004 ◽  
Vol 70 (12) ◽  
pp. 7413-7417 ◽  
Author(s):  
Mikael Courbot ◽  
Laurent Diez ◽  
Roberta Ruotolo ◽  
Michel Chalot ◽  
Pierre Leroy
Keyword(s):  
Molecular Mass ◽  
Ectomycorrhizal Fungi ◽  
High Performance ◽  
Metal Tolerance ◽  
Ectomycorrhizal Fungus ◽  
Paxillus Involutus ◽  
Cellular Mechanisms ◽  
Chromatography Method ◽  
Relative Lack ◽  
Liquid Chromatography Method

ABSTRACT Molecular and cellular mechanisms underlying the sustained metal tolerance of ectomycorrhizal fungi are largely unknown. Some of the main mechanisms involved in metal detoxification appear to involve the chelation of metal ions in the cytosol with thiol-containing compounds, such as glutathione, phytochelatins, or metallothioneins. We used an improved high-performance liquid chromatography method for the simultaneous measurement of thiol-containing compounds from cysteine and its derivatives (γ-glutamylcysteine, glutathione) to higher-molecular-mass compounds (phytochelatins). We found that glutathione and γ-glutamylcysteine contents increased when the ectomycorrhizal fungus Paxillus involutus was exposed to cadmium. An additional compound with a 3-kDa molecular mass, most probably related to a metallothionein, increased drastically in mycelia exposed to cadmium. The relative lack of phytochelatins and the presence of a putative metallothionein suggest that ectomycorrhizal fungi may use a different means to tolerate heavy metals, such as Cd, than do their plant hosts.

scholarly journals Genetic diversity study in Hungarian coldblooded horses

2017 ◽  
pp. 29-34
Author(s):  
Nikolett Csizmár ◽  
Sándor Mihók ◽  
András Jávor ◽  
Szilvia Kusza
Keyword(s):  
High Performance ◽  
Nucleotide Diversity ◽  
Base Pairs ◽  
Loop Region ◽  
Horse Breeding ◽  
D Loop ◽  
Hungarian Population ◽  
Genetic Diversity Study ◽  
Feeding Technology ◽  
Diversity Study

Because of the feeding technology innovation, accelerated transport and communication facilities breeds of high performance breeds replaced local autochone breeds worldwide. These latter species however have an important role in gene conservation. Hungarian cold-blooded horse breeding stock are lacking pedigree, the actual founder breed mares are not known. For this reason, it is an major priority defining the genetic backround of the existing flock, for that breeding could operate with purposeful using of origin maternal founders. In the present study 195 cold-blooded Hungarian mares tail and mane sample were analized. Our analysis was carried out between 15531–15752 base pairs in mithocrondial DNA D-loop region, which reported a total of 222 base pairs. Fourtyone polymorphic sites were determined, which resulted in 39 haplotypes (h=39). The average pairwise differences were k=6.825. High haplotype and nucleotide diversity values were observed (Hd=0.968±0.003, π=0.026±0.003). Based on the previously defined variable positions of haplotypes defined by Jansen et al (2002), we groupped our haplotypes into haplogroups. 23 percent of the studied population (45 mares) belonged to haplogroup F1. Nearly 97% of the analyzed population was classified into one of eight  haplogroups defined by Jansen.et al. (2002). This study gives genetic information nearly 25% of the Hungarian population. Another possibility would be patterning more mares or involving more genetic marker in the study which will assuming the possibility of a more comprehensive analysis.

scholarly journals Purification and characterization of a thermostable ADP-glucose pyrophosphorylase from Thermus caldophilus GK-24

1996 ◽  
Vol 319 (3) ◽  
pp. 977-983 ◽  
Author(s):  
Jeong Heon KO ◽  
Cheorl Ho KIM ◽  
Dae-Sil LEE ◽  
Yu Sam KIM
Keyword(s):  
Molecular Mass ◽  
Specific Activity ◽  
Gel Filtration ◽  
Higher Plant ◽  
Sds Page ◽  
Thermus Caldophilus ◽  
Internal Peptide ◽  
Adp Glucose Pyrophosphorylase ◽  
Fructose 1,6 Bisphosphate

An extremely thermostable ADP-glucose pyrophosphorylase (AGPase) has been purified from Thermus caldophilus GK-24 to homogeneity by chromatographic methods, including gel filtration and ion-exchange and affinity chromatography. The specific activity of the enzyme was enriched 134.8-fold with a recovery of 10.5%. The purified enzyme was a single band by SDS/PAGE with a molecular mass of 52 kDa. The homotetrameric structure of the native enzyme was determined by gel filtration analysis, which showed a molecular mass of 230 kDa on a Superose-12 column, indicating that the structure of the enzyme is different from the heterotetrameric structures of higher-plant AGPases. The enzyme was most active at pH 6.0. The activity was maximal at 73–78 °C and its half-life was 30 min at 95 °C. Kinetic and regulatory properties were characterized. It was found that AGPase activity could be stimulated by a number of glycolytic intermediates. Fructose 6-phosphate, fructose 1,6-bisphosphate, phenylglyoxal and glucose 6-phosphate were effective activators, of which fructose 1,6-bisphosphate was the most effective. The enzyme was inhibited by phosphate, AMP or ADP. ATP and glucose 1-phosphate gave hyperbolic-shaped rate-concentration curves in the presence or absence of activator. A remarkable aspect of the amino acid composition was the existence of the hydrophobic and Ala+Gly residues. The N-terminal and internal peptide sequences were determined and compared with known sequences of various sources. It was apparently similar to those of AGPases from other bacterial and plant sources, suggesting that the enzymes are structurally related.

Human serum and plasma have different sources of epidermal growth factor

1990 ◽  
Vol 259 (3) ◽  
pp. R545-R548 ◽  
Author(s):  
A. Lev-Ran ◽  
D. L. Hwang ◽  
D. S. Snyder
Keyword(s):  
Bone Marrow ◽  
Platelet Count ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Molecular Mass ◽  
High Performance ◽  
Regression Line ◽  
Bone Marrow Ablation ◽  
Epidermal Growth ◽  
Marrow Ablation

Epidermal growth factor (EGF) was determined by radioimmunoassay in serum, plasma, and urine of 23 patients undergoing ablative therapy followed by bone marrow transplantation. The difference between the serum and plasma values reflected the amount of EGF released from the platelets at the time of blood coagulation. Platelet-derived EGF strongly correlated with platelet count (r + 0.850, P less than 0.0001), and the intercept of the regression line was very close to zero; one platelet contained approximately 2.5 x 10(-18) g EGF. Correspondingly, when the platelet count dropped after bone marrow ablation from 222 +/- 97 to 33 +/- 13 x 10(9)/l, the serum EGF decreased from 603 +/- 222 to 65 +/- 41 pg/ml (P less than 0.0001). Plasma EGF content did not correlate with the platelet count and did not change significantly after bone marrow ablation (before and after the ablation, correspondingly, 290 +/- 80 and 332 +/- 99 pg/ml, P = 0.194). High-performance liquid chromatographic fractionation of serum and plasma showed different molecular mass distribution of EGF-immunoreactive fractions. The main molecular mass components of the plasma EGF did not change after bone marrow ablation. Urine excretion remained unchanged (320 +/- 133 and 314 +/- 173 pmol EGF/mmol creatinine). We conclude that whereas platelets are the source of serum EGF, the origin of plasma EGF is different and the search of its origin is warranted.

scholarly journals Collagenolytic Serine-Carboxyl Proteinase from Alicyclobacillus sendaiensis Strain NTAP-1: Purification, Characterization, Gene Cloning, and Heterologous Expression

2003 ◽  
Vol 69 (1) ◽  
pp. 162-169 ◽  
Author(s):  
Naoki Tsuruoka ◽  
Toru Nakayama ◽  
Masako Ashida ◽  
Hisashi Hemmi ◽  
Masahiro Nakao ◽  
...  
Keyword(s):  
Heterologous Expression ◽  
Molecular Mass ◽  
Gene Cloning ◽  
Collagen Type I ◽  
Protein Sequencing ◽  
Collagenolytic Activity ◽  
Efficient Production ◽  
Type I ◽  
Calculated Molecular Mass ◽  
Collagen Peptides

ABSTRACT Enzymatic degradation of collagen produces peptides, the collagen peptides, which show a variety of bioactivities of industrial interest. Alicyclobacillus sendaiensis strain NTAP-1, a slightly thermophilic, acidophilic bacterium, extracellularly produces a novel thermostable collagenolytic activity, which exhibits its optimum at the acidic region (pH 3.9) and is potentially applicable to the efficient production of such peptides. Here, we describe the purification to homogeneity, characterization, gene cloning, and heterologous expression of this enzyme, which we call ScpA. Purified ScpA is a monomeric, pepstatin-insensitive carboxyl proteinase with a molecular mass of 37 kDa which exhibited the highest reactivity toward collagen (type I, from a bovine Achilles tendon) among the macromolecular substrates examined. On the basis of the sequences of the peptides obtained by digestion of collagen with ScpA, the following synthetic peptides were designed as substrates for ScpA and kinetically analyzed: Phe-Gly-Pro-Ala*Gly-Pro-Ile-Gly (k cat, 5.41 s−1; Km , 32 μM) and Met-Gly-Pro-Arg*Gly-Phe-Pro-Gly-Ser (k cat, 351 s−1; Km , 214 μM), where the asterisks denote the scissile bonds. The cloned scpA gene encoded a protein of 553 amino acids with a calculated molecular mass of 57,167 Da. Heterologous expression of the scpA gene in the Escherichia coli cells yielded a mature 37-kDa species after a two-step proteolytic cleavage of the precursor protein. Sequencing of the scpA gene revealed that ScpA was a collagenolytic member of the serine-carboxyl proteinase family (the S53 family according to the MEROPS database), which is a recently identified proteinase family on the basis of crystallography results. Unexpectedly, ScpA was highly similar to a member of this family, kumamolysin, whose specificity toward macromolecular substrates has not been defined.

Ectoparasitic chiggers (Acari: Trombiculidae, Leeuwenhoekiidae), lice (Phthiraptera), and Hemiptera (Cimicidae and Reduviidae) from South Carolina, U.S.A.

Zootaxa ◽  
2004 ◽  
Vol 647 (1) ◽  
pp. 1 ◽  
Author(s):  
WILL K. REEVES ◽  
LANCE A. DURDEN ◽  
WILLIAM J. WRENN
Keyword(s):  
South Carolina ◽  
Trypanosoma Cruzi ◽  
Blood Feeding ◽  
Domestic Animals ◽  
The State ◽  
Cimex Lectularius ◽  
Intermediate Hosts ◽  
Bed Bug ◽  
Animal Pathogens

We report on the distribution of 15 chiggers, 31 lice of mammals, and 7 blood feeding hemipteran species in South Carolina. Some of these arthropods are vectors of pathogens to humans and domestic animals. Both Triatoma lecticularia and T. sanguisuga were reported from houses and these bugs are potential vectors of Trypanosoma cruzi. We also report on the continued presence of the bed bug, Cimex lectularius, in homes across the state. In addition we found the lice Haematopinus suis, Neohaematopinus sciuropteri, Pediculis humanus, Polyplax spinulosa, and Trichodectes canis, all of which are vectors or intermediate hosts of human or animal pathogens.

scholarly journals Lack of influence by endosymbiont Wolbachia on virus titer in the common bed bug, Cimex lectularius

2019 ◽  
Vol 12 (1) ◽  
Author(s):  
Michael L. Fisher ◽  
Jay F. Levine ◽  
James S. Guy ◽  
Hiroyuki Mochizuki ◽  
Matthew Breen ◽  
...  
Keyword(s):  
Blood Meal ◽  
Virus Titer ◽  
Blood Feeding ◽  
Cimex Lectularius ◽  
Future Research ◽  
Bed Bugs ◽  
Bed Bug ◽  
Post Feeding ◽  
Significant Difference ◽  
The Common

Abstract Background The common bed bug, Cimex lectularius, is an obligatory blood-feeding ectoparasite that requires a blood meal to molt and produce eggs. Their frequent biting to obtain blood meals and intimate association with humans increase the potential for disease transmission. However, despite more than 100 years of inquiry into bed bugs as potential disease vectors, they still have not been conclusively linked to any pathogen or disease. This ecological niche is extraordinarily rare, given that nearly every other blood-feeding arthropod is associated with some type of human or zoonotic disease. Bed bugs rely on the bacteria Wolbachia as an obligate endosymbiont to biosynthesize B vitamins, since they acquire a nutritionally deficient diet, but it is unknown if Wolbachia confers additional benefits to its bed bug host. In some insects, Wolbachia induces resistance to viruses such as Dengue, Chikungunya, West Nile, Drosophila C and Zika, and primes the insect immune system in other blood-feeding insects. Wolbachia might have evolved a similar role in its mutualistic association with the bed bug. In this study, we evaluated the influence of Wolbachia on virus replication within C. lectularius. Methods We used feline calicivirus as a model pathogen. We fed 40 bed bugs from an established line of Wolbachia-cured and a line of Wolbachia-positive C. lectularius a virus-laden blood meal, and quantified the amount of virus over five time intervals post-feeding. The antibiotic rifampicin was used to cure bed bugs of Wolbachia. Results There was a significant effect of time post-feeding, as the amount of virus declined by ~90% over 10 days in both groups, but no significant difference in virus titer was observed between the Wolbachia-positive and Wolbachia-cured groups. Conclusions These findings suggest that other mechanisms are involved in virus suppression within bed bugs, independent of the influence of Wolbachia, and our conclusions underscore the need for future research.

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