Uptake of proteins and degradation of human serum albumin by Plasmodium falciparum – infected human erythrocytes

BACKGROUND: Intraerythrocytic malaria parasites actively import obligate nutrients from serum and export proteins and lipids to erythrocyte cytoplasm and membrane. The import of macromolecules in the malaria parasite has been the subject of many debates. To understand the import of macromolecules by the parasite, we studied the uptake of proteins by Plasmodium falciparum infected human erythrocyte. METHODS: Proteins were biotin labelled individually, purified on a gel filtration column and added to uninfected and infected asynchronized culture. The uptake of these proteins by malaria parasites was determined by western blot analysis of parasite pellet and their different fractions using streptavidin-horseradish conjugate. To further confirm this import, we studied the uptake of (125)I-labelled proteins by western blot analysis as well as used direct immunofluorescence method. RESULTS: Here we show that biotin labelled and radio-iodinated polypeptides of molecular sizes in the range of 45 to 206 kDa, when added in the culture medium, get direct access to the parasite membrane through a membrane network by by-passing the erythrocyte cytosol. The import of these polypeptides is ATP-dependent as sodium azide treatment blocks this uptake. We also show that malaria parasites have the ability to take up and degrade biotin labelled human serum albumin, which has been shown to be essential for the parasite growth. CONCLUSIONS: These results can be used, as a basis to explore the role of human serum albumin in the intraerythrocytic development of parasites, and this in turn can be an important adjunct to the development of novel antimalarial drugs

MicroRNAs as Future Treatment Tools and Diagnostic Biomarkers in Alzheimer’s Disease

Alzheimer’s disease (AD) is a neurodegenerative disorder and is considered to be the most common form of dementia. This disorder is characterized by the formation of amyloid β (Aβ) plaques, neurofibrillary tangles, and alterations in synaptic function, all of which cause memory loss and behavioral disturbances. Despite the high prevalence of AD, effective therapeutic and diagnostic tools remain unavailable. MicroRNAs (miRNAs, miRs) are regulatory non-coding RNAs that target mRNAs. MiRNAs are involved in the regulation of the expressions of APP and BACE1, Aβ clearance, and the formation of neuro-fibrillary tangles. Furthermore, there are evidences that show alteration in the expression of several miRs in AD. MicroRNA is emerging as a biomarker because they have high specificity and, efficiency, and can be detected in biological fluids such as cerebrospinal fluid, tear, urine, blood. Moreover, miRNAs may be acquired and measured easily by utilizing real-time PCR, next-generation sequencing, or microarray. These techniques are cost-effective in comparison with imaging techniques such as magnetic resonance imaging, positron emission tomography. These features make miRNAs viable therapeutic as well as diagnostic tools in the treatment of AD. This review covers the regulatory function of miRNAs in AD, as well as their prospective applications as diagnostic biomarkers

Characterization of protective epitopes in a highly conserved Plasmodium falciparum antigenic protein containing repeats of acidic and basic residues

The delineation of putatively protective and immunogenic epitopes in vaccine candidate proteins constitutes a major research effort towards the development of an effective malaria vaccine. By virtue of its role in the formation of the immune clusters of merozoites, its location on the surface of merozoites, and its highly conserved nature both at the nucleotide sequence level and the amino acid sequence level, the antigen which contains repeats of acidic and basic residues (ABRA) of the human malaria parasite Plasmodium falciparum represents such an antigen. Based upon the predicted amino acid sequence of ABRA, we synthesized eight peptides, with six of these (AB-1 to AB-6) ranging from 12 to 18 residues covering the most hydrophilic regions of the protein, and two more peptides (AB-7 and AB-8) representing its repetitive sequences. We found that all eight constructs bound an appreciable amount of antibody in sera from a large proportion of P. falciparum malaria patients; two of these peptides (AB-1 and AB-3) also elicited a strong proliferation response in peripheral blood mononuclear cells from all 11 human subjects recovering from malaria. When used as carrier-free immunogens, six peptides induced a strong, boostable, immunoglobulin G-type antibody response in rabbits, indicating the presence of both B-cell determinants and T-helper-cell epitopes in these six constructs. These antibodies specifically cross-reacted with the parasite protein(s) in an immunoblot and in an immunofluorescence assay. In another immunoblot, rabbit antipeptide sera also recognized recombinant fragments of ABRA expressed in bacteria. More significantly, rabbit antibodies against two constructs (AB-1 and AB-5) inhibited the merozoite reinvasion of human erythrocytes in vitro up to ~90%. These results favor further studies so as to determine possible inclusion of these two constructs in a multicomponent subunit vaccine against asexual blood stages of P. falciparum

Characterization of protective epitopes in a highly conserved Plasmodium falciparum antigenic protein containing repeats of acidic and basic residues

The delineation of putatively protective and immunogenic epitopes in vaccine candidate proteins constitutes a major research effort towards the development of an effective malaria vaccine. By virtue of its role in the formation of the immune clusters of merozoites, its location on the surface of merozoites, and its highly conserved nature both at the nucleotide sequence level and the amino acid sequence level, the antigen which contains repeats of acidic and basic residues (ABRA) of the human malaria parasite Plasmodium falciparum represents such an antigen. Based upon the predicted amino acid sequence of ABRA, we synthesized eight peptides, with six of these (AB-1 to AB-6) ranging from 12 to 18 residues covering the most hydrophilic regions of the protein, and two more peptides (AB-7 and AB-8) representing its repetitive sequences. We found that all eight constructs bound an appreciable amount of antibody in sera from a large proportion of P. falciparum malaria patients; two of these peptides (AB-1 and AB-3) also elicited a strong proliferation response in peripheral blood mononuclear cells from all 11 human subjects recovering from malaria. When used as carrier-free immunogens, six peptides induced a strong, boostable, immunoglobulin G-type antibody response in rabbits, indicating the presence of both B-cell determinants and T-helper-cell epitopes in these six constructs. These antibodies specifically cross-reacted with the parasite protein(s) in an immunoblot and in an immunofluorescence assay. In another immunoblot, rabbit antipeptide sera also recognized recombinant fragments of ABRA expressed in bacteria. More significantly, rabbit antibodies against two constructs (AB-1 and AB-5) inhibited the merozoite reinvasion of human erythrocytes in vitro up to ∼90%. These results favor further studies so as to determine possible inclusion of these two constructs in a multicomponent subunit vaccine against asexual blood stages of P. falciparum

Transcriptional analysis of an immune-responsive serine protease from Indian malarial vector, Anopheles culicifacies

<p>Abstract</p> <p>Background</p> <p>The main vector for transmission of malaria in India is the <it>Anopheles culicifacies </it>mosquito species, a naturally selected subgroup of which is completely refractory (R) to transmission of the malaria parasite, <it>Plasmodium vivax</it>;</p> <p>Results</p> <p>Here, we report the molecular characterization of a serine protease (<it>acsp30</it>)-encoding gene from <it>A. culicifacies</it>, which was expressed in high abundance in the refractory strain compared to the susceptible (S) strain. The transcriptional upregulation of <it>acsp30 </it>upon <it>Plasmodium </it>challenge in the refractory strain coincided with ookinete invasion of mosquito midgut. Gene organization and primary sequence of <it>acsp30 </it>were identical in the R and S strains suggesting a divergent regulatory status of <it>acsp30 </it>in these strains. To examine this further, the upstream regulatory sequences of <it>acsp30 </it>were isolated, cloned and evaluated for the presence of promoter activity. The 702 bp upstream region of <it>acsp30 </it>from the two strains revealed sequence divergence. The promoter activity measured by luciferase-based reporter assay was shown to be 1.5-fold higher in the R strain than in the S. Gel shift experiments demonstrated a differential recruitment of nuclear proteins to upstream sequences of <it>acsp30 </it>as well as a difference in the composition of nuclear proteins in the two strains, both of which might contribute to the relative abundance of <it>acsp30 </it>in the R strain;</p> <p>Conclusion</p> <p>The specific upregulation of <it>acsp30 </it>in the R strain only in response to <it>Plasmodium </it>infection is suggestive of its role in contributing the refractory phenotype to the <it>A. culicifacies </it>mosquito population.</p

Identification of karyopherin β as an immunogenic antigen of the malaria parasite using immune mice and human sera

A differential immunoscreening of the λgt11 Plasmodium falciparum genomic expression library was carried out using anti-P. yoelii sera (convalescent-phase mouse sera) and immune sera collected from healthy adults, to identify novel cross-reactive and possibly protective antigens of the parasite. One clone, with an insert size of 1132 bp that reacted strongly with both the sera was selected. The insert was found to be a part of the P. falciparum karyopherin β (PfKβ) homologue. RT-PCR and Northern blot analysis confirmed the expression of PfKβ in the blood stages of the parasite. The ~110 kDa protein was localized in the cytoplasm at the ring and trophozoite, and in the parasitophorous vacuole at the schizont stage. Two large fragments of PfKβ representing the N- and C-terminal halves were expressed in E. coli. The recombinant proteins were highly immunogenic in mice, and also found to be the target for immune response in natural infections of Plasmodium spp. Anti-sera against the protein showed a low level of anti-parasitic activity. Immunization with recombinant PfKβ fragments was only partially protective against a heterologous challenge infection in mice. Our results show that the parasite releases a highly immunogenic, cytoplasmic protein into the host which may not contribute to the development of protective immunity

Electromagnetic interference shielding performance of carbon nanostructure reinforced, 3D printed polymer composites

We report the electrical, mechanical and electromagnetic interference (EMI) shielding performance of polypropylene random copolymer (PPR)/multi-wall carbon nanotube (MWCNT) nanocomposites enabled via customized fused filament fabrication process. The electro-conductive PPR/MWCNT filament feedstocks were fabricated via shear-induced melt-blending process that allows 3D printing of nanoengineered composites even at higher MWCNT loading (up to 8 wt%). The uniform dispersion of MWCNTs in PPR matrix confirmed via Raman spectroscopy and scanning electron microscopy facilitates better mechanical, electrical and EMI shielding performance. The results furthermore show enhanced shielding properties and higher attenuation for the nanocomposites printed in 90° direction (~ − 37 dB for 8 wt% MWCNT loading). Effective interfacial adhesion between the beads with lesser extent of voids (confirmed via micro-computed tomography) endorsed low transmission loss in nanocomposites printed in 90° direction compared to samples printed in 0° direction. Surface architected structure (frustum shape) reveals higher specific shielding effectiveness (maximum ~ − 40 dBg−1cm3, + 38%) over the plain structure. The realization of excellent shielding effectiveness (~ 99.9% attenuation) of additive manufacturing-enabled PPR/MWCNT nanocomposites demonstrates their potential for lightweight and strong EMI shields

Clinical outcome, viral response and safety profile of chloroquine in COVID-19 patients — initial experience

Introduction: Chloroquine and its analogues are currently being investigated for the treatment and post exposure prophylaxis of COVID-19 due to its antiviral activity and immunomodulatory activity.Material and methods: Confirmed symptomatic cases of COVID-19 were included in the study. Patients were supposed to receive chloroquine (CQ) 500 mg twice daily for 7 days. Due to a change in institutional protocol, initial patients received chloroquine and subsequent patients who did not receive chloroquine served as negative controls. Clinical effectiveness was determined in terms of timing of symptom resolution and conversion rate of reverse transcriptase polymerase chain reaction (RT-PCR) on day 14 and day 15 of admission.Results: Twelve COVID-19 patients formed the treatment arm and 17 patients were included in the control arm. The duration of symptoms among the CQ treated group (6.3 ± 2.7 days) was significantly (p-value = 0.009) lower than that of the control group (8.9 ± 2.2 days). There was no significant difference in the rate of RT-PCR negativity in both groups. 2 patients out of 12 developed diarrhea in the CQ therapy arm.  Conclusion: The duration of symptoms among the treated group (with chloroquine) was significantly lower than that of the control group. RT-PCR conversion was not significantly different between the 2 groups

B cell profiles, antibody repertoire and reactivity reveal dysregulated responses with autoimmune features in melanoma

B cells are known to contribute to the anti-tumor immune response, especially in immunogenic tumors such as melanoma, yet humoral immunity has not been characterized in these cancers to detail. Here we show comprehensive phenotyping in samples of circulating and tumor-resident B cells as well as serum antibodies in melanoma patients. Memory B cells are enriched in tumors compared to blood in paired samples and feature distinct antibody repertoires, linked to specific isotypes. Tumor-associated B cells undergo clonal expansion, class switch recombination, somatic hypermutation and receptor revision. Compared with blood, tumor-associated B cells produce antibodies with proportionally higher levels of unproductive sequences and distinct complementarity determining region 3 properties. The observed features are signs of affinity maturation and polyreactivity and suggest an active and aberrant autoimmune-like reaction in the tumor microenvironment. Consistent with this, tumor-derived antibodies are polyreactive and characterized by autoantigen recognition. Serum antibodies show reactivity to antigens attributed to autoimmune diseases and cancer, and their levels are higher in patients with active disease compared to post-resection state. Our findings thus reveal B cell lineage dysregulation with distinct antibody repertoire and specificity, alongside clonally-expanded tumor-infiltrating B cells with autoimmune-like features, shaping the humoral immune response in melanoma

A bibliography of parasites and diseases of marine and freshwater fishes of India

With the increasing demand for fish as human food, aquaculture both in freshwater and salt water is rapidly developing over the world. In the developing countries, fishes are being raised as food. In many countries fish farming is a very important economic activity. The most recent branch, mariculture, has shown advances in raising fishes in brackish, estuarine and bay waters, in which marine, anadromous and catadromous fishes have successfully been grown and maintained