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Abstract Background HCFC1 encodes transcriptional co‐regulator HCF‐1, which undergoes an unusual proteolytic maturation at a centrally located proteolysis domain. HCFC1 variants were associated with X‐linked cobalamin metabolism disorders and mental retardation‐3. This study aimed to explore the role of HCFC1 variants in common epilepsy and the mechanism underlying phenotype heterogeneity. Methods Whole‐exome sequencing was performed in a cohort of 313 patients with idiopathic partial (focal) epilepsy. Functional studies determined the effects of the variants on the proteolytic maturation of HCF‐1, cell proliferation and MMACHC expression. The role of HCFC1 variants in partial epilepsy was validated in another cohort from multiple centers. Results We identified seven hemizygous HCFC1 variants in 11 cases and confirmed the finding in the validation cohort with additional 13 cases and six more hemizygous variants. All patients showed partial epilepsies with favorable outcome. None of them had cobalamin disorders. Functional studies demonstrated that the variants in the proteolysis domain impaired the maturation by disrupting the cleavage process with loss of inhibition of cell growth but did not affect MMACHC expression that was associated with cobalamin disorder. The degree of functional impairment was correlated with the severity of phenotype. Further analysis demonstrated that variants within the proteolysis domain were associated with common and mild partial epilepsy, whereas those in the kelch domain were associated with cobalamin disorder featured by severe and even fatal epileptic encephalopathy, and those in the basic and acidic domains were associated with mainly intellectual disability. Conclusion HCFC1 is potentially a candidate gene for common partial epilepsy with distinct underlying mechanism of proteolysis dysfunction. The HCF‐1 domains played distinct functional roles and were associated with different clinical phenotypes, suggesting a sub‐molecular effect. The distinct difference between cobalamin disorders and idiopathic partial epilepsy in phenotype and pathogenic mechanism, implied a clinical significance in early diagnosis and management.

Cryptosporidium parvum is one of the major species causing mild to severe cryptosporidiosis in humans and animals. We have previously observed that 2-deoxy-d-glucose (2DG) could inhibit both the enzyme activity of C. parvum hexokinase (CpHK) and the parasite growth in vitro. However, the action and fate of 2DG in C. parvum was not fully investigated. In the present study, we showed that, although 2DG could be phosphorylated by CpHK to form 2DG-6-phosphate (2DG6P), the anti-cryptosporidial activity of 2DG was mainly attributed to the action of 2DG on CpHK, rather than the action of 2DG or 2DG6P on the downstream enzyme glucose-6-phosphate isomerase (CpGPI) nor 2DG6P on CpHK. These observations further supported the hypothesis that CpHK could serve as a drug target in the parasite. We also screened 1,200 small molecules consisting of marketed drugs against CpHK, from which four drugs were identified as CpHK inhibitors with micromolar level of anti-cryptospordial activities at concentrations nontoxic to the host cells (i.e. hexachlorphene, thimerosal, alexidine dihydrochloride, and ebselen with EC 50  = 0.53, 1.77, 8.1 and 165 μM, respectively). The anti-CpHK activity of the four existing drugs provided us new reagents for studying the enzyme properties of the parasite hexokinase., (© 2018 International Society of Protistologists.)

We developed nested PCR protocols and performed a multiyear survey on the prevalence of several protozoan parasites in wild northern bobwhite (Colinus virginianus) and scaled quail (Callipepla squamata) in the Rolling Plains ecoregion of Texas and Oklahoma (i.e. fecal pellets, bird intestines and blood smears collected between 2010 and 2013). Coccidia, cryptosporidia, and microsporidia were detected in 46.2%, 11.7%, and 44.0% of the samples (n = 687), whereas histomona and hematozoa were undetected. Coccidia consisted of one major and two minor Eimeria species. Cryptosporidia were represented by a major unknown Cryptosporidium species and Cryptosporidium baileyi. Detected microsporidia species were highly diverse, in which only 11% were native avian parasites including Encephalitozoon hellem and Encephalitozoon cuniculi, whereas 33% were closely related to species from insects (e.g. Antonospora, Liebermannia, and Sporanauta). This survey suggests that coccidia infections are a significant risk factor in the health of wild quail while cryptosporidia and microsporidia may be much less significant than coccidiosis. In addition, the presence of E. hellem and E. cuniculi (known to cause opportunistic infections in humans) suggests that wild quail could serve as a reservoir for human microsporidian pathogens, and individuals with compromised or weakened immunity should probably take precautions while directly handling wild quail., (© 2016 The Author(s) Journal of Eukaryotic Microbiology © 2016 International Society of Protistologists.)

Cryptosporidium parvum is unable to synthesize fatty acids de novo, but possesses three long-chain fatty acyl-CoA synthetase (CpACS) isoforms for activating fatty acids. We have recently shown that these enzymes could be targeted to kill the parasite in vitro and in vivo. Here, we demonstrated that the CpACS genes were differentially expressed during the parasite life cycle, and their proteins were localized to different subcellular structures by immunofluorescence and immuno-electron microscopies. Among them, CpACS1 displayed as an apical protein in sporozoites and merozoites, but no or little presence during the intracellular merogony until the release of merozoites, suggesting that CpACS1 probably functioned mainly during the parasite invasion and/or early stage of intracellular development. Both CpACS2 and CpACS3 proteins were present in all parasite life cycle stages, in which CpACS2 was present in the parasite and the parasitophorous vacuole membranes (PVM), whereas CpACS3 was mainly present in the parasite plasma membranes with little presence in the PVM. These observations suggest that CpACS2 and CpACS3 may participate in scavenging and transport of fatty acids across the PVM and the parasite cytoplasmic membranes, respectively., (© 2015 The Author(s) Journal of Eukaryotic Microbiology © 2015 International Society of Protistologists.)

We have cloned and sequenced a lactate dehydrogenase (LDH) gene from Cryptosporidium parvum (CpLDH1). With this addition, and that of four recently deposited alpha-proteobacterial malate dehydrogenase (MDH) genes, the phylogenetic relationships among apicomplexan LDH and bacterial MDH were re-examined. Consistent with previous studies, our maximum likelihood (ML) analysis using the quartet-puzzling method divided 105 LDH/MDH enzymes into five clades, and confirmed that mitochondrial MDH is a sister clade to those of y-proteobacteria, rather than to alpha-proteobacteria. In addition, a Cryptosporidium parvum MDH (CpMDH1) was identified from the ongoing Cryptosporidium genome project that appears to belong to a distinct clade (III) comprised of 22 sequences from one archaebacterium, numerous eubacteria, and several apicomplexans. Using the ML puzzling test and bootstrapping analysis with protein distance and parsimony methods, the resulting trees not only robustly confirmed the alpha-proteobacterial relationship of apicomplexan LDH/MDH, but also supported a monophyletic relationship of CpLDH1 with CpMDHI. These data suggest that, unlike most other eukaryotes, the Apicomplexa may be one of the few lineages retaining an alpha-proteobacterial-type MDH that could have been acquired from an ancestral alpha-proteobacterium through primary endosymbiosis giving rise to the mitochondria, or through an unknown lateral gene transfer (LGT) event.