A machine learning approach to identify hydrogenosomal proteins in Trichomonas vaginalis

The protozoan parasite Trichomonas vaginalis is the causative agent of trichomoniasis, the most widespread nonviral sexually transmitted disease in humans. It possesses hydrogenosomes-anaerobic mitochondria that generate H(2), CO(2), and acetate from pyruvate while converting ADP to ATP via substrate-level phosphorylation. T. vaginalis hydrogenosomes lack a genome and translation machinery; hence, they import all their proteins from the cytosol. To date, however, only 30 imported proteins have been shown to localize to the organelle. A total of 226 nuclear-encoded proteins inferred from the genome sequence harbor a characteristic short N-terminal presequence, reminiscent of mitochondrial targeting peptides, which is thought to mediate hydrogenosomal targeting. Recent studies suggest, however, that the presequences might be less important than previously thought. We sought to identify new hydrogenosomal proteins within the 59,672 annotated open reading frames (ORFs) of T. vaginalis, independent of the N-terminal targeting signal, using a machine learning approach. Our training set included 57 gene and protein features determined for all 30 known hydrogenosomal proteins and 576 nonhydrogenosomal proteins. Several classifiers were trained on this set to yield an import score for all proteins encoded by T. vaginalis ORFs, predicting the likelihood of hydrogenosomal localization. The machine learning results were tested through immunofluorescence assay and immunodetection in isolated cell fractions of 14 protein predictions using hemagglutinin constructs expressed under the homologous SCSα promoter in transiently transformed T. vaginalis cells. Localization of 6 of the 10 top predicted hydrogenosome-localized proteins was confirmed, and two of these were found to lack an obvious N-terminal targeting signal.     

Characterization of Prototheca wickerhamii Isolated from Disseminated Algaemia of Kidney Transplant Patient from Malaysia

Prototheca wickerhamii isolated from blood of 61-year-old kidney transplant patient was described. Although it is classified as an alga (genus Chlorella), the disease, protothecosis, is included under mycoses because of its clinical pathological presentations. Colony characteristics of P. wickerhamii are indistinguishable from other yeast-like organisms like Cryptococcus and Candida. Fortunately, commercial identification system for yeast can be used to identify this organism to the species level. Electron microscopy demonstrated "morula" or daisy-like appearance of its endosporulating sporangia. The organism was sensitive to amphotericin B by E test method. Even though human protothecosis is uncommon, it cannot be ignored because it is emerging as an opportunistic infection in immunosuppressed individuals. To our knowledge, this is the first reported case of disseminated algaemia due to P. wickerhamii in Malaysia.     

Rotationally oscillating drill (Ros-Drill) for mouse ICSI without using mercury

Intracytoplasmic sperm injection (ICSI) is an important assisted reproductive technology (ART). Due to deployment difficulties and low efficiency of the earlier (conventional) version of ICSI, especially in the mouse, a piezo-assisted ICSI technique had evolved as a popular ART methodology in recent years. An important and remaining problem with this technique, however, is that it requires small amounts of mercury to stabilize the pipette tip when piezoelectric force pulses are applied. To eliminate this problem we developed and tested a completely different and mercury-free technology, called the "Ros-Drill" (rotationally oscillating drill). The technique uses microprocessor-controlled rotational oscillations on a spiked micropipette without mercury or piezo. Preliminary experimental results show that this new microinjection technology gives high survival rate (>70% of the injected oocytes) and fertilization rate (>80% of the survived oocytes), and blastocyst formation rates in early trials (approximately 50% of the survived oocytes). Blastocysts created by Ros-Drill ICSI were transferred into the uteruses of pseudopregnant surrogate mothers and healthy pups were born and weaned. The Ros-Drill ICSI technique is automated and therefore; it requires a very short preliminary training for the specialists, as evidenced in many successful biological trials. These advantages of Ros-Drill ICSI over conventional and piezo-assisted ICSI are clearly demonstrated and it appears to have resolved an important problem in reproductive biology.     

In Vitro Antifungal Susceptibility of <Emphasis Type="Italic">Neoscytalidium dimidiatum</Emphasis> Clinical Isolates from Malaysia

Neoscytalidium dimidiatum is an opportunistic fungus causing cutaneous infections mostly, which are difficult to treat due to antifungal resistance. In Malaysia, N. dimidiatum is associated with skin and nail infections, especially in the elderly. These infections may be mistaken for dermatophyte infections due to similar clinical appearance. In this study, Neoscytalidium isolates from cutaneous specimens, identified using morphological and molecular methods (28 Neoscytalidium dimidiatum and 1 Neoscytalidium sp.), were evaluated for susceptibility towards antifungal agents using the CLSI broth microdilution (M38-A2) and Etest methods. Amphotericin B, voriconazole, miconazole and clotrimazole showed high in vitro activity against all isolates with MIC ranging from 0.0313 to 1 µg/mL. Susceptibility towards fluconazole and itraconazole was noted in up to 10% of isolates, while ketoconazole was inactive against all isolates. Clinical breakpoints for antifungal drugs are not yet available for most filamentous fungi, including Neoscytalidium species. However, the results indicate that clinical isolates of N. dimidiatum in Malaysia were sensitive towards miconazole, clotrimazole, voriconazole and amphotericin B, in vitro.     

Exponential Spread of Hepatitis C Virus Genotype 4a in Egypt

Hepatitis C virus (HCV) infects >10% of the general population in Egypt, in which intravenous injection with an antimony compound for endemic schistosomiasis in the past has been implicated. To simulate the epidemic history of HCV in Egypt, sera were obtained from 3608 blood donors at 13 governorates in or surrounding the Nile valley during 1999. The prevalence of antibody to HCV (anti-HCV) and genotypes was determined in them, and the molecular evolutionary analysis based on the neutral theory was applied to HCV isolates of genotype 4a, which is outstandingly prevalent in Egypt and indigenous there. Of 3608 sera, 317 (8.8%) were positive for anti-HCV. The molecular evolutionary analysis on 47 HCV genotype 4a isolates of carriers from various districts in Egypt indicated that the spread of HCV-4a would have increased exponentially during the 1940s through 1980 when oral medications became available. In conclusion, the estimated spread time is consistent with the duration of intravenous antimony campaigns in Egypt.     

In vitro effect of carbonic anhydrase inhibitor acetazolamide on cell viability,migration and colony formation of colorectal cancer cells

Acidification of extracellular medium in malignant tumors increases the invasive behaviors of cancer cells. In normal healthy tissues, acid production is catalyzed by carbonic anhydrases. Some of the carbonic anhydrase enzymes are overexpressed in certain types of cancer. The present study aimed to investigate the effect of acetazolamide, a potent carbonic anhydrase inhibitor, on in vitro cultivated cancer cells. Three different assays (MTT test, wound healing and clonogenic assay) were performed using human colorectal adenocarcinoma cells (SW620) to evaluate the suppressive effect of acetazolamide, on the colorectal cancer cells migration ability, colony formation and cell viability. The dose-dependent (1–1000 μM) reducing effect of acetazolamide on the cell viability was more significant within the first 48 h. This inhibitory effect of acetazolamide was found to be decreased at 72 h, and affects cells migration ability of cells at 24 and 48 h. Acetazolamide was observed to inhibit the cell viability, migration and colony formation ability of cells, depending on dose.     

Mixed Herbs Drugs: Inhibitory Effect on Growth of the Endogenous Mycoflora and Aflatoxin Production

Twenty commercial mixed herbal drugs were examined for mycological profile. Aspergillus species were the predominant fungi found in the drugs. Other fungi harboured in the drugs with less frequency were Paecilomyces species, Eurotium species, Monascus species, Acremonium species, Penicillium species, Cladosporium species, Scopulariopsis species, Phialophora species and Fonseceae species. Fungal count was between 1.0 log₁₀ CFU and 2.4 log₁₀ CFU per gram of sample. When the drugs were incubated in 85% humidity at 25°C, fungal colonies grew on only two of the drugs. The mixed herbal drugs were extracted with water and the extracts were used to grow Aspergillus parasiticus. All extracts reduced aflatoxin B₁ and aflatoxin G₁ production by 62–97%. All but two of the extracts reduced aflatoxin B₂ and aflatoxin G₂ production by 39–95%. It can be concluded that the commercial powdered mixed herbal drugs contained low number of endogenous fungi, and these drugs are inhibitory to the growth of its endogenous fungi and aflatoxins production by aflatoxigenic fungi.     

The effects of extracorporeal shock wave lithotripsy on antioxidant enzymes in erythrocytes

The effects of extracorporeal shock wave lithotripsy (ESWL) on patients undergoing ESWL for renal stone treatment have been studied using activities of glucose-6-phosphate dehydrogenase (G6PDH), superoxide dismutase (SOD) and catalase (CAT), and levels of malondialdehyde (MDA) in the erythrocyte haemolysate. The study included 23 patients (eight women, 15 men with an age range of 23-57 years). Blood samples were taken 5 min before ESWL, in addition to 1 h and 5 days after termination of treatment. Enzyme activities and MDA levels in erythrocytes were measured spectrophotometrically. When compared with the values obtained before ESWL, erythrocyte G6PDH (p = 0.015), SOD (p = 0.036) and CAT (p = 0.01) activities were found to be significantly reduced at the first hour after ESWL. On the fifth day after ESWL, erythrocyte enzyme activities were normalized to the values obtained before ESWL. Although there was a significant difference between values before and 1 h after ESWL (p = 0.003), no difference was detected between 1 h after ESWL and 5 days after ESWL (p > 0.05) in terms of MDA values. The findings of the present study revealed that erythrocyte lipid peroxidation might be induced and antioxidative defence mechanism may be transiently impaired by ESWL.