scholarly journals Molecular Characterization of Annexin B2, B3 and B12 in Taenia multiceps

Genes ◽  
2018 ◽  
Vol 9 (11) ◽  
pp. 559 ◽  
Author(s):  
Cheng Guo ◽  
Yue Xie ◽  
Yuchen Liu ◽  
Ning Wang ◽  
Jiafei Zhan ◽  
...  
Keyword(s):  
Developmental Stages ◽  
Polyclonal Antibodies ◽  
Parasite Development ◽  
Economic Losses ◽  
Mrna Levels ◽  
Western Blots ◽  
Taenia Multiceps ◽  
Homologous Genes ◽  
Coenurus Cerebralis

Coenurus cerebralis, the metacestode of Taenia multiceps, causes coenurosis, a disease severely affecting goat, sheep, cattle and yak farming and resulting in huge economic losses annually. Annexins bind calcium ions and play an important role in flatworm parasite development. To explore potential functions of annexins in T. multiceps, three homologous genes, namely, TmAnxB2, TmAnxB3 and TmAnxB12, were screened from the transcriptome dataset, amplified from C. cerebralis cDNA and subjected to bioinformatics analysis. Then, polyclonal antibodies recognizing the recombinant TmAnxB2 (rTmAnxB2) and rTmAnxB3 were prepared for localization of TmAnxB2 and TmAnxB3 in different tissues and developmental stages by immunofluorescence. The transcription of all three genes was also measured by relative fluorescent quantitative PCR. The sizes of rTmAnxB2, rTmAnxB3 and rTmAnxB12 were 58.00, 53.06 and 53.51 kDa, respectively, and rTmAnxB12 was unstable. Both rTmAnxB2 and rTmAnxB3 were recognized by goat-positive T. multiceps sera in Western blots. Immunofluorescence revealed that TmAnxB2 and TmAnxB3 were localized in the protoscolex and cyst wall and TmAnxB3 was also detected in adult cortex. TmAnxB2 and TmAnxB12 mRNA levels were determined to be highest in oncospheres and protoscolex, whereas transcription of TmAnxB3 was highest in scolex and immature segments. Taken together, these findings indicate that TmAnxB2 and TmAnxB12 may play critical roles in T. multiceps larvae, while TmAnxB3 may have important functions in adults. These results will lay the foundation for functional research of annexins in T. multiceps.

Ultrastructural and molecular characterization of Bacillidium vesiculoformis n. sp. (Microspora: Mrazekiidae) in the freshwater oligochaete Nais simplex (Oligochaeta: Naididae)

Parasitology ◽  
2004 ◽  
Vol 130 (1) ◽  
pp. 31-40 ◽  
Author(s):  
D. J. MORRIS ◽  
R. S. TERRY ◽  
K. B. FERGUSON ◽  
J. E. SMITH ◽  
A. ADAMS
Keyword(s):  
Phylogenetic Analysis ◽  
New Species ◽  
Infected Cell ◽  
Host Cell ◽  
Developmental Stages ◽  
Parasite Development ◽  
Adhesive Quality ◽  
Transmission Studies ◽  
A New Species

The development of a new species, Bacillidium vesiculoformis n. sp. (Microspora, Mrazekiidae), is described from the freshwater oligochaete Nais simplex (Oligochaeta, Naididae). Initial stages of parasite development consist of a monokaryotic merogony within a haemocyte of the intestinal blood sinus. The resulting hypertrophied haemocyte is attached to the chloragocytes of the sinus by fine cytoplasmic extensions with the sinus around the cell becoming greatly enlarged. The meronts within the haemocyte form diplokaryotic sporonts that undergo sporogenesis directly within the cytoplasm of the host cell. The infected cell becomes packed with spores and developmental stages, causing it dramatically to increase in size, eventually rupturing the oligochaete and cell. Sporogony appears to be disporoblastic. Released spores were observed to have an adhesive quality. Transmission studies conducted with mature spores failed to transmit the parasite horizontally although vertical transmission was observed. Phylogenetic analysis of the parasite demonstrated that B. vesiculoformis clustered with microsporidian parasites of bryozoa and two other microsporidians, Janacekia debaiseuxi and an unidentified Bacillidium sp.

scholarly journals Characterization of an MLP Homologue from Haemaphysalis longicornis (Acari: Ixodidae) Ticks

Pathogens ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 284
Author(s):  
Jin Luo ◽  
Hui Shen ◽  
Qiaoyun Ren ◽  
Guiquan Guan ◽  
Bo Zhao ◽  
...  
Keyword(s):  
Muscle Development ◽  
Developmental Stages ◽  
Rabbit Antiserum ◽  
Haemaphysalis Longicornis ◽  
Mrna Levels ◽  
Muscle Lim Protein ◽  
Rapid Amplification ◽  
Differentiation And Proliferation ◽  
Phosphate Buffered Saline

Members of the cysteine-rich protein (CRP) family are known to participate in muscle development in vertebrates. Muscle LIM protein (MLP) belongs to the CRP family and has an important function in the differentiation and proliferation of muscle cells. In this study, the full-length cDNA encoding MLP from Haemaphysalis longicornis (H. longicornis; HLMLP) ticks was obtained by 5′ rapid amplification of cDNA ends (RACE). To verify the transcriptional status of MLP in ticks, HLMLP gene expression was assessed during various developmental stages by real-time PCR (RT-PCR). Interestingly, HLMLP expression in the integument was significantly (P < 0.01) higher than that observed in other tested tissues of engorged adult ticks. In addition, HLMLP mRNA levels were significantly downregulated in response to thermal stress at 4 °C for 48 h. Furthermore, recombinant HLMLP was expressed in Escherichia coli, and Western blot analysis showed that rabbit antiserum against H. longicornis adults recognized HLMLP and MLPs from different ticks. Ten 3-month-old rabbits that had never been exposed to ticks were used for the immunization and challenge experiments. The rabbits were divided into two groups of five rabbits each, where rabbits in the first group were immunized with HLMLP, while those in the second group were immunized with phosphate-buffered saline (PBS) diluent as controls. The vaccination of rabbits with the recombinant HLMLP conferred partial protective immunity against ticks, resulting in 20.00% mortality and a 17.44% reduction in the engorgement weight of adult ticks. These results suggest that HLMLP is not ideal as a candidate for use in anti-tick vaccines. However, the results of this study generated novel information on the MLP gene in H. longicornis and provide a basis for further investigation of the function of this gene that could potentially lead to a better understanding of the mechanism of myofiber determination and transformation

Molecular characterization and phylogenetic analysis of Taenia multiceps from China

2018 ◽  
Vol 63 (4) ◽  
pp. 721-727 ◽  
Author(s):  
L. Tan ◽  
A.B. Wang ◽  
S.Q. Zheng ◽  
X.L. Zhang ◽  
C.J. Huang ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Dna Sequences ◽  
Hunan Province ◽  
Biological Study ◽  
Economic Losses ◽  
Molecular Characteristics ◽  
12S Rrna ◽  
Intermediate Hosts ◽  
Taenia Multiceps ◽  
Coenurus Cerebralis

Abstract Taenia multiceps, one of the most widely distributed zoonotic tapeworm parasites, is able to parasitize the small intestine of canids. The metacestode of T.multiceps is fatal to ruminants and causes important economic losses in livestock. However, molecular characteristics of T.multiceps and coenurus in China are still unclear. In this study, 36 goat isolates of the coenurus stage and 18 dog isolates of the adult stage of T.multiceps were obtained from three geographical areas in China and the isolated parasite above were analyzed by amplifying the partial of cytochrome coxidase subunit 1(pcox1), 12S ribosomal RNA (12S rRNA) from mitochondrial DNA (mtDNA) regions and an internal transcribed spacer (ITS) of ribosomal DNA (rDNA). These DNA sequences obtained from T.multiceps and coenurus were employed to evaluate the nucleotide diversity and confirm the relationship between T.multiceps and coenurus. Sequences variation were 0–1.4%, 0–1.5%, 0–4.2% for pcox1, 12S rRNA and ITS, respectively, among T.multiceps and coenurus isolates obtained in this study. In Sichuan province, sequence variations for Coenurus cerebralis isolated from Yaan city were 0–1.4% for pcox1, 0–1.0% for 12S rRNA and 0–2.1% for ITS. In Hunan province, variations were 0–1.0%, 0–1.5% and 0–3.3% for corresponding genes for non-coenurus cerebralis isolated from Changsha city, while variations of T.multiceps isolates from Xiangxi autonomous prefecture were 0–1.0%, 0–1.1% and 0–3.4% for pcox1, 12S rRNA and ITS, respectively. Phylogenetic analysis based on pcox1 sequences indicated that all cerebral and noncerebral metacestodes belong to T.multiceps. These results provide reference values for future molecular epidemiological and biological study on T.multiceps in dogs and intermediate hosts.

scholarly journals The Production and characterization of Polyclonal Antibodies Against Interferon Alpha in Mice

2021 ◽  
Vol 3 (4) ◽  
Author(s):  
Mehreen Fatima ◽  
Fatima Khalid ◽  
Azra Quraishi
Keyword(s):  
Immune Response ◽  
Autoimmune Diseases ◽  
Interferon Alpha ◽  
Polyclonal Antibodies ◽  
Mouse Serum ◽  
Interferon Production ◽  
Dot Blot ◽  
Western Blots ◽  
Response Activation

The polyclonal antibodies are used extensively for research purposes in many areas of biology, such as immunoprecipitation, histochemistry, enzyme linked immunosorbent assays (ELISA), diagnosis of disease and western blots. Typically, an animal’s immune system will generate a large group of antibodies that recognize several epitopes of a particular antigen. Interferon alpha plays an important role in immune response activation and therefore is of interest in studies related to autoimmune diseases. In this paper the production of antibodies against interferon was studied in order to quantify interferon production to analyze interferon levels in autoimmune disorders in the future. For the antibody production, one month old laboratory grade mice were injected with interferon alpha in combination with a Freund’s complete adjuvant for a course of five weeks after which the antibodies were obtained in mouse serum. Confirmation of the production of anti-interferon alpha antibodies was carried out by the Elisa, immune dot blot and western blot analysis. An interferon alpha of approximately 20.5-21.5 KDa was detected in immunedot blot test.  These antibodies may be produced in these mouse models commercially and could be used in future for treatment of autoimmune diseases by managing the interferon levels in the patients. Copyright(c) The Authors

Identification and characterization of a novel specific secreted protein family for selected members of the subfamily Ostertagiinae (Nematoda)

Parasitology ◽  
2007 ◽  
Vol 135 (1) ◽  
pp. 63-70 ◽  
Author(s):  
H. SAVERWYNS ◽  
A. VISSER ◽  
A. J. NISBET ◽  
I. PEELAERS ◽  
K. GEVAERT ◽  
...  
Keyword(s):  
Developmental Stages ◽  
Cell Damage ◽  
Inflammatory Cells ◽  
Glycosylation Site ◽  
Protein Family ◽  
Protein Motif ◽  
Western Blots ◽  
Teladorsagia Circumcincta ◽  
Identification And Characterization

SUMMARYIt has been shown that the bovine abomasal parasite, Ostertagia ostertagi, drastically modulates its microenvironment, causing epithelial cell damage, accumulation of inflammatory cells and pH changes in the stomach. The mechanisms used by the parasite to change the abomasal environment are largely unknown, but an important role has been attributed to excretory-secretory (ES) products from the parasite. In this study we have identified proteins representing a novel ES protein family, characterized by the SCP/Tpx-1/Ag5/PR-1/Sc7 protein motif. These proteins were named Oo-AL1 and Oo-AL2 (O. ostertagi ASP-like protein). Both proteins contain a signal peptide and 1 predicted N-glycosylation site. The transcript for Oo-AL1 was present from the L4 stage onwards in both male and female adult worms, whereas the Oo-AL2 transcript was hardly detectable. Western blots of somatic extracts and ES products from different developmental stages of O. ostertagi, probed with anti-Oo-AL1 antibodies, revealed Oo-AL proteins in the ES products of adult worms. An analysis of the nematode genome and EST databases indicated that these novel ES proteins are unique to O. ostertagi and its relative, Teladorsagia circumcincta, suggesting a key function in these abomasal parasites.

Molecular detection and immunological localization of gill Na+/H+ exchanger in the dogfish (Squalus acanthias)

2008 ◽  
Vol 294 (3) ◽  
pp. R1092-R1102 ◽  
Author(s):  
James B. Claiborne ◽  
Keith P. Choe ◽  
Alison I. Morrison-Shetlar ◽  
Jill C. Weakley ◽  
Justin Havird ◽  
...  
Keyword(s):  
Polyclonal Antibodies ◽  
Squalus Acanthias ◽  
Transport Systems ◽  
Rectal Gland ◽  
Mrna Levels ◽  
Western Blots ◽  
Reading Frame ◽  
Antisense Probe ◽  
Acid Secreting

The dogfish ( Squalus acanthias) can make rapid adjustments to gill acid-base transfers to compensate for internal acidosis/alkalosis. Branchial Na+/H+ exchange (NHE) has been postulated as one mechanism driving the excretion of H+ following acidosis. We have cloned gill cDNA that includes an open reading frame coding for a 770-residue protein most homologous (∼71%) to mammalian NHE2. RT-PCR revealed NHE2 transcripts predominantly in gill, stomach, rectal gland, intestine, and kidney. In situ hybridization with an antisense probe against NHE2 in gill sections revealed a strong mRNA signal from a subset of interlamellar and lamellae cells. We developed dogfish-specific polyclonal antibodies against NHE2 that detected a ∼70-kDa protein in Western blots and immunologically recognized branchial cells having two patterns of protein expression. Cytoplasmic and apical NHE2 immunoreactivity were observed in cells coexpressing basolateral Na+-K+-ATPase. Other large ovoid cells more generally staining for NHE2 also were strongly positive for basolateral H+-ATPase. Gill mRNA levels for NHE2 and H+-ATPase did not change following systemic acidosis (as measured by quantitative PCR 2 h after a 1- or 2-meq/kg acid infusion). These data indicate that posttranslational adjustments of NHE2 and other transport systems (e.g., NHE3) following acidosis may be of importance in the short-term pH adjustment and net branchial H+ efflux observed in vivo. NHE2 may play multiple roles in the gills, involved with H+ efflux from acid-secreting cells, basolateral H+ reabsorption for pHi regulation, and in parallel with H+-ATPase for the generation of HCO3− in base-secreting cells.

Molecular characterization of mitofilin (HMP), a mitochondria-associated protein with predicted coiled coil and intermembrane space targeting domains

1996 ◽  
Vol 109 (9) ◽  
pp. 2253-2264 ◽  
Author(s):  
P.R. Odgren ◽  
G. Toukatly ◽  
P.L. Bangs ◽  
R. Gilmore ◽  
E.G. Fey
Keyword(s):  
Polyclonal Antibodies ◽  
Limited Proteolysis ◽  
Coiled Coil ◽  
Cell Types ◽  
Cdna Clones ◽  
Intermembrane Space ◽  
Western Blots ◽  
Double Label ◽  
Helical Region

We have identified and characterized a human protein of the mitochondria which we call mitofilin. Using monoclonal and polyclonal antibodies, we have isolated cDNA clones and characterized mitofilin biochemically. It appears as a 90 and 91 kDa doublet in western blots and is translated from a single 2.7 kb mRNA. Antibodies raised against cellular and bacterially-expressed protein given identical cytoplasmic immunofluorescence and immunoblot results. Mitofilin co-localizes with mitochondria in immunofluorescence experiments and co-purifies with mitochondria. Double label studies show co-localization only with mitochondria and not with Golgi or endoplasmic reticulum. Co-localization with mitochondria is retained when actin or tubulin are de-polymerized, and mitofilin is expressed in all human cell types tested. The cDNA encodes a polypeptide with a central alpha-helical region with predicted coiled coil domains flanked by globular amino and carboxy termini. Unlike coiled coil motor proteins, mitofilin is resistant to detergent extraction. The presence of mitochondrial targeting and stop-transfer sequences, along with the accessibility of mitofilin to limited proteolysis suggests that it resides predominantly in the intermembrane space, consistent with immuno-electron micrographs which show mitofilin mainly at the mitochondrial periphery. The cDNA sequence of mitofilin is identical to that recently reported by Icho et al. (1994; Gene 144, 301–306) for a mRNA preferentially expressed in heart muscle (HMP), consistent with the high levels of mitochondria in cardiac myocytes.

scholarly journals Comparative Transcriptome Analyses of the Developmental Stages of Taenia multiceps

2021 ◽  
Vol 8 ◽  
Author(s):  
Wen-Hui Li ◽  
Yang Yang ◽  
Nian-Zhang Zhang ◽  
Jian-Kui Wang ◽  
Yin-Ju Liu ◽  
...  
Keyword(s):  
Information Processing ◽  
Rna Sequencing ◽  
Developmental Stages ◽  
Central Nervous System Disease ◽  
Read Length ◽  
Taenia Multiceps ◽  
System Disease ◽  
Host Interaction ◽  
Coenurus Cerebralis ◽  
Cerebral Coenurosis

Cerebral coenurosis, caused by the larvae of Taenia multiceps (Coenurus cerebralis), is a fatal central nervous system disease in sheep and other herbivores and occasionally humans. Comparative transcriptomic profiles of the developmental stages of the parasite remain unknown. In this study, RNA sequencing was used to determine the transcriptome profiles of different stages of the life cycle of T. multiceps, including Oncosphere, Coenurus cerebralis (Pro with Cyst), and Adult (Adu), as well as scolex-neck proglottids (Snp), immature–mature proglottids (Imp), and gravid proglottids (Grp) of the adult stage. A total of 42.6 Gb (average 6.1 Gb) Illumina pair-end reads with a 125-bp read length were generated for seven samples. The total number of differentially expressed genes (DEGs) in the various life stages ranged from 2,577 to 3,879; however, for the tissues of the adult worm, the range was from 1,229 to 1,939. Kyoto Encyclopedia of Genes and Genomes analysis showed that the DEGs mainly participated in cellular and metabolic processes, binding and catalytic activity, genetic information processing, and environmental information processing. In addition, a large number of genes related to development and parasite–host interaction were identified. Quantitative reverse transcription-polymerase chain reaction confirmed that the levels of 28 selected DEGs were consistent with those determined using RNA sequencing. The present study provides insights into the mechanisms of the development and parasitic life of T. multiceps.

Characterization of putative membrane protein genes of the ‘Candidatus Phytoplasma asteris’, chrysanthemum yellows isolate

2008 ◽  
Vol 54 (5) ◽  
pp. 341-351 ◽  
Author(s):  
Luciana Galetto ◽  
Jacqueline Fletcher ◽  
Domenico Bosco ◽  
Massimo Turina ◽  
Astri Wayadande ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Expression Vector ◽  
Polyclonal Antibodies ◽  
Western Blots ◽  
Protein Fragments ◽  
Aster Yellows ◽  
Euscelidius Variegatus ◽  
Macrosteles Quadripunctulatus ◽  
Aster Yellows Phytoplasma

To characterize potentially important surface-exposed proteins of the phytoplasma causing chrysanthemum yellows (CY), new primers were designed based on the conserved regions of 3 membrane protein genes of the completely sequenced onion yellows and aster yellows witches’ broom phytoplasmas and were used to amplify CY DNA. The CY genes secY, amp, and artI, encoding the protein translocase subunit SecY, the antigenic membrane protein Amp and the arginine transporter ArtI, respectively, were cloned and completely sequenced. Alignment of CY-specific secY sequences with the corresponding genes of other phytoplasmas confirmed the 16S rDNA-based classification, while amp sequences were highly variable within the ‘Candidatus Phytoplasma asteris’. Five CY partial sequences were cloned into the pRSetC expression vector, and 3 of the encoded protein fragments (Amp 64/651, Amp 64/224, ArtI 131/512) were expressed as fusion antigens for the production of CY-specific polyclonal antibodies (A416 against Amp 64/224; A407 against ArtI 131/512). A416 recognized, in Western blots, the full-length Amp from CY-infected plants (periwinkle, daisy) and insect vectors ( Euscelidius variegatus , Macrosteles quadripunctulatus ). A416 also reacted to European aster yellows, to primula yellows phytoplasmas, to northern Italian strains of ‘Ca. Phytoplasma asteris’ from lettuce and gladiolus, but it did not react to American aster yellows phytoplasma.

Synthesis of gill Na+-K+-ATPase in Atlantic salmon smolts: differences in α-mRNA and α-protein levels

2000 ◽  
Vol 278 (1) ◽  
pp. R101-R110 ◽  
Author(s):  
Helena D'Cotta ◽  
Claudiane Valotaire ◽  
Florence le Gac ◽  
Patrick Prunet
Keyword(s):  
Atlantic Salmon ◽  
Atpase Activity ◽  
Early Stage ◽  
Polyclonal Antibodies ◽  
Mrna Abundance ◽  
Activity Measurement ◽  
Mrna Levels ◽  
Western Blots ◽  
Α Subunit ◽  
Protein Levels

Several parameters were analyzed to determine the mechanisms responsible for the enhancement of the gill Na+-K+-ATPase activity of Atlantic salmon smolts. A major α-subunit transcript of 3.7 kb was revealed by Northern blot in both parr and smolt gills when hybridized with two distinct cDNA probes. The α-mRNA abundance demonstrated an increase to maximal levels in smolts at an early stage of the parr-smolt transformation. This was followed by a gradual rise in α-protein levels, revealed by Western blots with specific antibodies and by an increase in gill Na+-K+-ATPase hydrolytic activity, both only reaching maximum levels a month later, at the peak of the transformation process. Parr fish experienced a decrease in α-mRNA abundance and had basal levels of α-protein and enzyme activity. Measurement of the binding of [3H]ouabain to Na+-K+-ATPase was characterized in smolts and parr gill membranes showing more than a twofold elevation in smolts and was of high affinity in both groups (dissociation constant = 20–23 nM). Modulation of the enzyme due to increased salinity was also observed in seawater-transferred smolts, as demonstrated by an increase in α-mRNA levels after 24 h with a rise in Na+-K+-ATPase activity occurring only after 11 days. No qualitative change in α-expression was revealed at either the mRNA or protein level. Immunological identification of the α-protein was performed with polyclonal antibodies directed against the rat α-specific isoforms, revealing that parr, freshwater, and seawater smolts have an α3-like isoform. This study shows that the increase in Na+-K+-ATPase activity in smolt gills depends first on an increase in the α-mRNA expression and is followed by a slower rise in α-protein abundance that eventually leads to a higher synthesis of Na+-K+pumps.

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