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The execution of the apoptotic death program in metazoans is characterized by a sequence of morphological and biochemical changes that include cell shrinkage, presentation of phosphatidylserine at the cell surface, mitochondrial alterations, chromatin condensation, nuclear fragmentation, membrane blebbing and the formation of apoptotic bodies. Methodologies for measuring apoptosis are based on these markers. Except for membrane blebbing and formation of apoptotic bodies, all other events have been observed in most protozoan parasites undergoing cell death. However, while techniques exist to detect these markers, they are often optimised for metazoan cells and therefore may not pick up subtle differences between the events occurring in unicellular organisms and multi-cellular organisms.In this review we discuss the markers most frequently used to analyze cell death in protozoan parasites, paying special attention to changes in cell morphology, mitochondrial activity, chromatin structure and plasma membrane structure/permeability. Regarding classical regulators/executors of apoptosis, we have reviewed the present knowledge of caspase-like and nuclease activities.

Binding at the interface: We tested the inhibitory activity of a set of peptide sequences derived from an α-helix of the dimeric trypanothione reductase from Leishmania infantum. Replacement of a glutamic acid residue with a lysine promoted monomer dissociation and enzyme inhibition.

No abstract

Biallelic loss-of-function (LOF) mutations of the NCF4 gene, encoding the p40 phox subunit of the phagocyte NADPH oxidase, have been described in only 1 patient. We report on 24 p40 phox -deficient patients from 12 additional families in 8 countries. These patients display 8 different in-frame or out-of-frame mutations of NCF4 that are homozygous in 11 of the families and compound heterozygous in another. When overexpressed in NB4 neutrophil-like cells and EBV-transformed B cells in vitro, the mutant alleles were found to be LOF, with the exception of the p.R58C and c.120_134del alleles, which were hypomorphic. Particle-induced NADPH oxidase activity was severely impaired in the patients' neutrophils, whereas PMAinduced dihydrorhodamine-1,2,3 (DHR) oxidation, which is widely used as a diagnostic test for chronic granulomatous disease (CGD), was normal or mildly impaired in the patients. Moreover, the NADPH oxidase activity of EBV-transformed B cells was also severely impaired, whereas that of mononuclear phagocytes was normal. Finally, the killing of Candida albicans and Aspergillus fumigatus hyphae by neutrophils was conserved in these patients, unlike in patients with CGD. The patients suffer from hyperinflammation and peripheral infections, but they do not have any of the invasive bacterial or fungal infections seen in CGD. Inherited p40 phox deficiency underlies a distinctive condition, resembling a mild, atypical form of CGD. phox -deficient patient is compound heterozygous for a premature stop codon and a missense mutation in the PX domain that compromises binding to PI(3)P, which results in the impairment of neutrophil phagocytosis-induced oxidase activity (26). As in classic CGD neutrophils, intracellular oxidant production after stimulation with serum-opsonized zymosan (SOZ), IgG beads, or serumopsonized Staphylococcus aureus is impaired in the patient's neutrophils, and S. aureus killing is also defective (20,26,30). However, in this patient, unlike in classic CGD patients, the production of O 2 -by neutrophils in response to stimulation with PMA or formyl-methionyl-leucyl-phenylalanine (fMLF) is normal (26). The killing of S. aureus by neutrophils is also impaired in p40 phox deficiency and classic CGD are largely unknown. Here, we describe the characteristics of 24 patients from 12 families in 8 countries with biallelic mutations of NCF4. The Journal of Clinical Investigation R E S E A R C H A R T I C L E

Cryptosporidium parvum and C. hominis are the most relevant species of this genus for human health. Both cause a self-limiting diarrhea in immunocompetent individuals, but cause potentially life-threatening disease in the immunocompromised. Despite the importance of these pathogens, only one reference genome of each has been analyzed and published. These two reference genomes were sequenced using automated capillary sequencing; as of yet, no next generation sequencing technology has been applied to improve their assemblies and annotations. For C. hominis, the main challenge that prevents a larger number of genomes to be sequenced is its resistance to axenic culture. In the present study, we employed next generation technology to analyse the genomic DNA and RNA to generate a new reference genome sequence of a C. hominis strain isolated directly from human stool and a new genome annotation of the C. parvum Iowa reference genome.

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Anopheles (Nyssorhynchus) benarrochi, An. (N.) oswaldoi, and An. (N.) rangeli are the most common anthropophilic mosquitoes in the southern Colombian state of Putumayo. Adult females are most commonly collected in epidemiological studies, and this stage poses significant problems for correct identification, due to overlapping inter-specific morphological characters. Although An. rangeli is easy to identify, the morphological variant of An. benarrochi found in the region and An. oswaldoi are not always easy to separate. Herein we provide a rapid molecular method to distinguish these two species in Southern Colombia. Sequence data for the second internal transcribed spacer (ITS2) region of rDNA was generated for link-reared progeny of An. benarrochi and An. oswaldoi, that had been identified using all life stages. ITS2 sequences were 540 bp in length in An. benarrochi (n = 9) and 531 bp in An. oswaldoi (n = 7). Sequences showed no intra-specific variation and ungapped inter-specific sequence divergence was 6.4%. Species diagnostic banding patterns were recovered following digestion of the ITS2 amplicons with the enzyme Hae III as follows: An. benarrochi (365, 137, and 38 bp) and An. oswaldoi (493 and 38 bp). This polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay provides rapid, accurate, and inexpensive species diagnosis of adult females. This will benefit future epidemiological studies and, as PCR amplification can be achieved using a single mosquito leg, the remaining specimen can be either retained as a morphological voucher or further used in vector incrimination studies. That An. benarrochi comprises a complex of at least two species across Latin America is discussed