Characterization of lymphocyte subsets in patients with common variable immunodeficiency reveals subsets of naive human B cells marked by CD24 expression

Increased proportions of naive B cell subset and B cells defined as CD27(neg)CD21(neg)CD38(neg) are frequently found in patients with common variable immunodeficiency (CVID) syndrome. Current methods of polychromatic flow cytometry and PCR-based detection of κ deletion excision circles allow for fine definitions and replication history mapping of infrequent B cell subsets. We have analyzed B cells from 48 patients with CVID and 49 healthy controls to examine phenotype, frequency, and proliferation history of naive B cell subsets. Consistent with previous studies, we have described two groups of patients with normal (CVID-21norm) or increased (CVID-21lo) proportions of CD27(neg)CD21(neg)CD38(neg) B cells. Upon further analyses, we found two discrete subpopulations of this subset based on the expression of CD24. The B cell subsets showed a markedly increased proliferation in CVID-21lo patients as compared with healthy controls, suggesting developmental arrest rather than increased bone marrow output. Furthermore, when we analyzed CD21(pos) naive B cells, we found two different subpopulations based on IgM and CD24 expression. They correspond to follicular (FO) I and FO II cells previously described in mice. FO I subset is significantly underrepresented in CVID-21lo patients. A comparison of the replication history of naive B cell subsets in CVID patients and healthy controls implies refined naive B cell developmental scheme, in which human transitional B cells develop into FO II and FO I. We propose that the CD27(neg)CD21(neg)CD38(neg) B cells increased in some of the CVID patients originate from the two FO subsets after loss of CD21 expression.     

Trypanosoma cruzi I genotypes in different geographical regions and transmission cycles based on a microsatellite motif of the intergenic spacer of spliced-leader genes

The intergenic region of spliced-leader (SL-IR) genes from 105 Trypanosoma cruzi I (Tc I) infected biological samples, culture isolates and stocks from 11 endemic countries, from Argentina to the USA were characterised, allowing identification of 76 genotypes with 54 polymorphic sites from 123 aligned sequences. On the basis of the microsatellite motif proposed by Herrera et al. (2007) to define four haplotypes in Colombia, we could classify these genotypes into four distinct Tc I SL-IR groups, three corresponding to the former haplotypes Ia (11 genotypes), Ib (11 genotypes) and Id (35 genotypes); and one novel group, Ie (19 genotypes). Genotypes harbouring the Tc Ic motif were not detected in our study. Tc Ia was associated with domestic cycles in southern and northern South America and sylvatic cycles in Central and North America. Tc Ib was found in all transmission cycles from Colombia. Tc Id was identified in all transmission cycles from Argentina and Colombia, including Chagas cardiomyopathy patients, sylvatic Brazilian samples and human cases from French Guiana, Panama and Venezuela. Tc Ie gathered five samples from domestic Triatoma infestans from northern Argentina, nine samples from wild Mepraia spinolai and Mepraia gajardoi and two chagasic patients from Chile and one from a Bolivian patient with chagasic reactivation. Mixed infections by Tc Ia + Tc Id, Tc Ia + Tc Ie and Tc Id + Tc Ie were detected in vector faeces and isolates from human and vector samples. In addition, Tc Ia and Tc Id were identified in different tissues from a heart transplanted Chagas cardiomyopathy patient with reactivation, denoting histotropism. Trypanosoma cruzi I SL-IR genotypes from parasites infecting Triatoma gerstaeckeri and Didelphis virginiana from USA, T. infestans from Paraguay, Rhodnius nasutus and Rhodnius neglectus from Brazil and M. spinolai and M. gajardoi from Chile are to our knowledge described for the first time.     

Studies on trimethoprim:hydroxypropyl-β-cyclodextrin: aggregate and complex formation

The present study is focused on the characterization of the interaction between trimethoprim, a dihydropteroate synthesase inhibitor, and hydroxypropyl-β-cyclodextrin (HP-β-CD) in aqueous solution and solid state. The freeze-drying method was used to prepare solid complexes, while simple blending was employed to obtain physical mixtures. The phase solubility was AN type, and demonstrated that trimethoprim solubility was significantly increased upon complexation with HP-β-CD. Conductivity experiments showed the presence of aggregates that explains the type profile for the solubility isotherm. The critical concentration for the aggregate formation was determined to be 69.3 mg/ml for pure HP-β-CD and 117.7 mg/ml in the presence of trimethoprim. Nuclear magnetic resonance spectroscopy provided evidence of trimethoprim:HP-β-CD molecular interaction in solution. Moreover, the complex was characterized in solid stated using Fourier-transform infrared spectroscopy (FT-IR) and scanning electron microscopy (SEM). The use of differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA) showed that the thermal stability of the drug is enhanced in the presence of HP-β-CD.     

Changes in macrophage membrane properties during early Leishmania amazonensis infection differ from those observed during established infection and are partially explained by phagocytosis

Understanding the impact of intracellular pathogens on the behavior of their host cells is key to designing new interventions. We are interested in how Leishmania alters the electrical function of the plasma membrane of the macrophage it infects. The specific question addressed here is the impact of Leishmania infection on macrophage membrane properties during the first 12 h post-infection. A decrease of 29% in macrophage membrane capacitance at 3 h post-infection indicates that the phagolysosome membrane is donated on entry by the macrophage plasma membrane. Macrophage membrane potential depolarized during the first 12 h post-infection, which associated with a decreased inward potassium current density, changed in inward rectifier conductance and increased outward potassium current density. Decreased membrane capacitance and membrane potential, with no changes in ion current density, were found in macrophages after phagocytosis of latex beads. Therefore we suggest that the macrophage membrane changes observed during early Leishmania infection appear to be associated with the phagocytic and activation processes.     

Comparisons of stress proteins and soluble carbohydrate in encysted embryos of Artemia franciscana and two species of Parartemia

We compared stress proteins (p26, artemin, hsp70) and alcohol-soluble carbohydrates (ASC) in cysts of Artemia franciscana and two as yet un-named species populations of Parartemia, the brine shrimp endemic to Australia. The small stress proteins and molecular chaperones, p26 and artemin, previously thought to be restricted to Artemia, and present in very large amounts in its encysted embryos (cysts), were also detected by western blotting in Parartemia cysts, even though roughly 85-100 million years have passed since these genera diverged. We interpret this finding as further evidence for the adaptive importance of these proteins in coping with the severe stresses these encysted embryos endure. As expected, hsp70 was present in all three groups of cysts, but apparently at somewhat lower concentrations in those of Parartemia. Based on measurements of ASC we propose that the disaccharide trehalose, critical for desiccation tolerance in many animal cells, has probably also been maintained in the metabolic repertoire of Parartemia whose cysts have well developed tolerance to severe desiccation.     

Lactobacillus casei administration reduces lung injuries in a Streptococcus pneumoniae infection in mice

The effect of the oral administration of Lactobacillus casei on the prevention of a Streptococcus pneumoniae lung infection in a mouse experimental model was studied, analyzing the innate and specific immune response. Adult Swiss albino mice were treated with L. casei (10(9)CFU/day) for 2, 5 and 7 d. Mice were infected intranasally with S. pneumoniae (10(6)CFU/mouse) after each treatment and the microbiological, histopathological and host responses were determined for 15 d after infection. Feeding L. casei for 2 d induced a faster clearance of S. pneumoniae, with a lower number of pneumococci in lung and a shorter period of septicemia than in the control group. L. casei administration induced activation of phagocytes as evidenced by the strong myeloperoxidase activity and the nitro blue tetrazolium assay in lung. Mice given L. casei for 2 d showed higher levels of anti-pneumococcal serum IgG and bronchoalveolar lavage IgA than the control mice. The group fed L. casei for 2 d could beneficially regulate the balance between tumor necrosis factor alpha and interleukin 10, allowing a more effective immune response against infection and modulating the inflammatory response, with less damage to the lung.     

Disruption of the endoplasmic reticulum and increases in cytoplasmic calcium are early events in cell death induced by the natural triterpenoid Asiatic acid

Triterpenoids are a novel class of compounds being investigated as potential therapeutic agents for the treatment of prostate cancer and other malignancies. Asiatic acid (AA) is a member of the ursane family of triterpenoids and has anticancer activity, but its mechanism of action is not completely understood. To investigate its mechanism of action, PPC-1 prostate cancer cells were treated with AA at increasing concentrations and times. AA induced rapid caspase-dependent and independent cell death that peaked within 8 h of treatment. AA-induced death was associated with early activation of caspases 2, 3, and 8, but not caspase 9. Within 2.5 h of treatment, release of calcium from intracellular stores and dilatation of the endoplasmic reticulum was observed. Thus, disruption of the endoplasmic reticulum and alterations in calcium homeostasis are early events in AA-induced death.     

First report of mites (Gamasida: Laelapidae) parasitic on wild rodents in Uruguay, with new host records

Information is presented for the first time on laelapid mites associated with wild rodents in Uruguay. Specimens of the following species were identified: Laelapinae: Androlaelaps fahrenholzi (Berlese), Androlaelaps rotundus (Fonseca), Gigantolaelaps wolffsohni (Oudemans), Laelaps paulistanensis (Fonseca), Laelaps manguinhosi (Fonseca) and Mysolaelaps microspinosus Fonseca; Haemogamasinae: Eulaelaps stabularis (Koch). Most of the ectoparasite-host associations are reported for the first time. New host and locality records presented in this study are in accordance with previous findings on the same and/or related host species in nearby South American localities.     

Hyperthermia reduces viral load of white spot syndrome virus in Penaeus vannamei

We have previously reported that white spot syndrome virus-infected Penaeus vannamei (also called Litopenaeus vannamei) maintained at 32 degrees C show higher survival rates and a significant increase in number of apoptotic cells when compared to infected shrimp kept at 26 degrees C. As apoptosis plays an important part in the antiviral response of invertebrates, we hypothesized that this process would reduce WSSV replication, allowing the shrimp to control the disease and survive. To test this hypothesis, shrimp were orally infected and maintained at either 26 degrees C (Group 1) or 32 degrees C (Group 2), DNA was extracted from haemolymph collected at various times from 6 to 216 h post-infection, and the number of viral units was quantified by real time PCR using SYBR Green. In parallel, histological examination was carried out to confirm the WSSV infection and to rule out concomitant diseases. Linear regression of real time PCR units (rtPCRU) of WSSV from Group 1 showed a significant increase with time post-infection (r2 = 0.7383; p < 0.001). Conversely, there was no increase in rtPCRU with time post-infection in Group 2 (r2 = 0.142), indicating that hyperthermia inhibited, either directly or indirectly, viral replication. In addition, comparison between the groups showed no difference in WSSV rtPCRU up to 48 h post-infection. After 72 h, shrimp from Group 1 had a significantly higher viral rtPCRU (ANOVA, p < 0.001). We conclude that hyperthermia-associated WSSV rtPCRU reduction could reflect either an increase in the shrimp antiviral response, or a direct negative effect on viral replication, or both.