Surface Area Loss and Increased Sphericity Account for the Splenic Entrapment of Subpopulations of Plasmodium falciparum Ring-Infected Erythrocytes

Ex vivo perfusion of human spleens revealed innate retention of numerous cultured Plasmodium falciparum ring-infected red blood cells (ring-iRBCs). Ring-iRBC retention was confirmed by a microsphiltration device, a microbead-based technology that mimics the mechanical filtering function of the human spleen. However, the cellular alterations underpinning this retention remain unclear. Here, we use ImageStream technology to analyze infected RBCs’ morphology and cell dimensions before and after fractionation with microsphiltration. Compared to fresh normal RBCs, the mean cell membrane surface area loss of trophozoite-iRBCs, ring-iRBCs and uninfected co-cultured RBCs (uRBCs) was 14.2% (range: 8.3–21.9%), 9.6% (7.3–12.2%) and 3.7% (0–8.4), respectively. Microsphilters retained 100%, ∼50% and 4% of trophozoite-iRBCs, ring-iRBCs and uRBCs, respectively. Retained ring-iRBCs display reduced surface area values (estimated mean, range: 17%, 15–18%), similar to the previously shown threshold of surface-deficient RBCs retention in the human spleen (surface area loss: >18%). By contrast, ring-iRBCs that successfully traversed microsphilters had minimal surface area loss and normal sphericity, suggesting that these parameters are determinants of their retention. To confirm this hypothesis, fresh normal RBCs were exposed to lysophosphatidylcholine to induce a controlled loss of surface area. This resulted in a dose-dependent retention in microsphilters, with complete retention occurring for RBCs displaying >14% surface area loss. Taken together, these data demonstrate that surface area loss and resultant increased sphericity drive ring-iRBC retention in microsphilters, and contribute to splenic entrapment of a subpopulation of ring-iRBCs. These findings trigger more interest in malaria research fields, including modeling of infection kinetics, estimation of parasite load, and analysis of risk factors for severe clinical forms. The determination of the threshold of splenic retention of ring-iRBCs has significant implications for diagnosis (spleen functionality) and drug treatment (screening of adjuvant therapy targeting ring-iRBCs).     

Mitochondrial Genome of the Eyeworm,Thelazia callipaeda (Nematoda: Spirurida), as the First Representative from the Family Thelaziidae

Human thelaziosis is an underestimated parasitic disease caused by Thelazia species (Spirurida: Thelaziidae). The oriental eyeworm, Thelazia callipaeda, infects a range of mammalian definitive hosts, including canids, felids and humans. Although this zoonotic parasite is of socio-economic significance in Asian countries, its genetics, epidemiology and biology are poorly understood. Mitochondrial (mt) DNA is known to provide useful genetic markers to underpin fundamental investigations, but no mt genome had been characterized for any members of the family Thelaziidae. In the present study, we sequenced and characterized the mt genome of T. callipaeda. This AT-rich (74.6%) mt genome (13,668 bp) is circular and contains 12 protein-coding genes, 22 transfer RNA genes and two ribosomal RNA genes, but lacks an atp8 gene. All protein-coding genes are transcribed in the same direction; the gene order is the same as those of Dirofilaria immitis and Setaria digitata (Onchocercidae), but distinct from Dracunculus medinensis (Dracunculidae) and Heliconema longissimum (Physalopteridae). Phylogenetic analyses of the concatenated amino acid sequence data for all 12 protein-coding genes by Bayesian inference (BI) showed that T. callipaeda (Thelaziidae) is related to the family Onchocercidae. This is the first mt genome of any member of the family Thelaziidae and should represent a new source of genetic markers for studying the epidemiology, ecology, population genetics and systematics of this parasite of humans and other mammals.     

Number and Function of Bone-Marrow Derived Angiogenic Cells and Coronary Flow Reserve in Women without Obstructive Coronary Artery Disease: A Substudy of the NHLBI-Sponsored Women's Ischemia Syndrome Evaluation (WISE)

Background

In women with ischemia and no obstructive coronary artery disease, the Women''s Ischemic Syndrome Evaluation (WISE) observed that microvascular coronary dysfunction (MCD) is the best independent predictor of adverse cardiovascular events. Since coronary microvascular tone is regulated in part by endothelium, we hypothesized that circulating endothelial cells (CEC), which reflect endothelial injury, and the number and function of bone-marrow derived angiogenic cells (BMDAC), which could help repair damaged endothelium, may serve as biomarkers for decreased coronary flow reserve (CFR) and MCD.

Methods

We studied 32 women from the WISE cohort. CFR measurements in response to intracoronary adenosine were taken as an index of MCD. We enumerated BMDAC colonies and CEC in peripheral blood samples. BMDAC function was assessed by assay of migration of CD34+ cells toward SDF-1 and measurement of bioavailable nitric oxide (NO). These findings were compared with a healthy reference group and also entered into a multivariable model with CFR as the dependent variable.

Results

Compared with a healthy reference group, women with MCD had lower numbers of BMDAC colonies [16 (0, 81) vs. 24 (14, 88); P = 0.01] and NO [936 (156, 1875) vs. 1168 (668, 1823); P = 0.02]. Multivariable regression analysis showed strong correlation of CFR to the combination of BMDAC colony count and CD34+ cell function (migration and NO) (R² = 0.45; P<0.05).

Conclusions

The BMDAC function and numbers of BMDAC colonies are decreased in symptomatic women with MCD and are independently associated with CFR. These circulating cells may provide mechanistic insights into MCD in women with ischemia.     

A Bacterial Phosphatase-Like Enzyme of the Malaria Parasite Plasmodium falciparum Possesses Tyrosine Phosphatase Activity and Is Implicated in the Regulation of Band 3 Dynamics during Parasite Invasion

Eukaryotic parasites of the genus Plasmodium cause malaria by invading and developing within host erythrocytes. Here, we demonstrate that PfShelph2, a gene product of Plasmodium falciparum that belongs to the Shewanella-like phosphatase (Shelph) subfamily, selectively hydrolyzes phosphotyrosine, as shown for other previously studied Shelph family members. In the extracellular merozoite stage, PfShelph2 localizes to vesicles that appear to be distinct from those of rhoptry, dense granule, or microneme organelles. During invasion, PfShelph2 is released from these vesicles and exported to the host erythrocyte. In vitro, PfShelph2 shows tyrosine phosphatase activity against the host erythrocyte protein Band 3, which is the most abundant tyrosine-phosphorylated species of the erythrocyte. During P. falciparum invasion, Band 3 undergoes dynamic and rapid clearance from the invasion junction within 1 to 2 s of parasite attachment to the erythrocyte. Release of Pfshelph2 occurs after clearance of Band 3 from the parasite-host cell interface and when the parasite is nearly or completely enclosed in the nascent vacuole. We propose a model in which the phosphatase modifies Band 3 in time to restore its interaction with the cytoskeleton and thus reestablishes the erythrocyte cytoskeletal network at the end of the invasion process.     

Influence of carbon and nitrogen sources on antifungal metabolite production by bacterium associated with entomopathogenic nematode against Penicillium expansum

A specific symbiotic Bacillus sp. isolated from a rhabditid entomopathogenic nematode, Rhabditis (Oscheius) sp. was found to produce large number of bioactive compounds. The present study was conducted to determine the effect of carbon and nitrogen sources for the production of antimicrobial substances by Bacillus sp. The yield of the crude antimicrobial substances and antimicrobial activity against the test micro-organism also differed significantly when carbon and nitrogen sources in the fermentation media were changed. The antifungal activity was significantly high in yeast extract plus fructose (46.5?±?2.12?mm) followed by yeast extract plus maltose, beef extract plus fructose and meat infusion plus glucose. High pressure liquid chromatography analysis of the crude antimicrobial substances revealed different peaks with different retention time indicating that they produced different compounds. When the carbon source was not included in the fermentation media, the antimicrobial production was substantially reduced. The results indicate that carbon source in the fermentation media plays a vital role in the production of antifungal substances. It is concluded that yeast extract and fructose as nitrogen and carbon sources produced maximum activity, which can effectively control the blue mould caused by Penicillium expansum in apples and pears.     

Plant virus infection development as affected by heavy metal stress

The work was focused on the investigation of possible dependencies between the development of viral infection in plants and the presence of high heavy metal concentrations in soil. Field experiments have been conducted in order to study the development of systemic tobacco mosaic virus (TMV) infection in Lycopersicon esculentum L. cv. Miliana plants under effect of separate salts of heavy metals Cu, Zn and Pb deposited in soil. As it is shown, simultaneous effect of viral infection and heavy metals in tenfold maximum permissible concentration leads to decrease of total chlorophyll content in experiment plants mainly due to the degradation of chlorophyll a. The reduction of chlorophyll concentration under the combined influence of both stress factors was more serious comparing to the separate effect of every single factor. Plants' treatment with toxic concentrations of lead and zinc leaded to slight delay in the development of systemic TMV infection together with more than twofold increase of virus content in plants that may be an evidence of synergism between these heavy metal's and virus' effects. Contrary, copper although decreased total chlorophyll content but showed protective properties and significantly reduced amount of virus in plants.     

Localization of human gene loci using spontaneous chromosome rearrangements in human-Chinese hamster somatic cell hybrids.

Analysis of human-Chinese hamster somatic cell hybrids with spontaneously derived chromosome structural changes has provided data for the regional and subregional localization of gene loci which have previously been assigned to human chromosomes 2, 12, and X. Correlation of the expression of human gene loci with the human chromosome complements present in somatic cell hybrids indicates that the cytoplasmic malate dehydrogenase (MDH1) locus is in the 2p23yields2pter region, and red cell acid phosphatase (AcP1) is at or adjacent to 2p23. The cytoplasmic isocitrate dehydrogenase (IDH1) locus is at or adjacent to 2q11, peptidase B (Pep B) is at or adjacent to 12q21, lactate dehydrogenase B (LDH B) is in the 12q21yiedls12pter region, glucose-6-phosphate dehydrogenase (G6PD) is in the Xq24yieldsXqter region, and the gene loci for phosphoglycerate kinase (PGK), alpha-galactosidase (alpha-gal), and hypoxanthine guanine phosphoribosyltransferase (GPRT) are in the Xp21yieldsXq24 region.     

Assignment of the αB-crystallin gene to human chromosome 11

Using a human αB-crystallin genomic probe and human-mouse somatic cell hybrids, the human αB-gene was assigned to chromosome 11 and further corroborated by in situ hybridization to normal metaphase chromosomes. This assignment confirmed and regionally mapped the locus to q22.3–23.1.     

Assignment of the human intestinal Na+/glucose cotransporter gene (SGLT1) to the q11.2----qter region of chromosome 22

The chromosomal location of the human intestinal Na+/glucose cotransporter gene (SGLT1) was determined using human cDNA and genomic probes for this transporter gene. Southern blot analysis of genomic DNA from 15 mouse-human somatic cell hybrids showed that the human gene for this transporter resides on chromosome 22. Analysis of hamster-human hybrids selectively retaining chromosome 22 or a portion of it allowed specific assignment of the locus to the q11.2----qter region of chromosome 22. A restriction fragment length polymorphism was identified with EcoRI.