Baseline TGF-ß correlates with protection after immunization with Plasmodium falciparum sporozoites in the controlled human malaria infection model.

BACKGROUND: Here we assessed a possible relationship between baseline TGF-ß concentrations and acquisition of sterile immunity after Plasmodium falciparum sporozoite immunization. METHODS: TGF-ß concentrations were determined in samples of 65 malaria-naive volunteers in 4 studies either prior to and after challenge infection, or prior to and after first immunizing infection under chemoprophylaxis with P. falciparum sporozoites. RESULTS: High baseline TGF-ß concentrations were associated with rapid acquisition of sterile protection (p = 0.028). CONCLUSION: Baseline TGF-ß concentrations predict the efficiency of acquisition of sterile immunity following sporozoite immunization and may represent a steady-state regulatory mechanism to keep in check immune systems with a low threshold for activation.

Vermamoeba vermiformis resides in water-based heater-cooler units and can enhance Mycobacterium chimaera survival after chlorine exposure.

BACKGROUND: Mycobacterium chimaera colonizes water-based heater-cooler units (HCUs), from which it can spread to patients during surgery. Vermamoeba vermiformis is a free-living waterborne amoeba, which was consistently present within HCUs. AIM: To determine whether these amoebae can be involved in the persistent presence of M. chimaera. METHODS: An in-vitro disinfection model. FINDINGS: Increased survival of M. chimaera was observed after chlorine exposure in the presence of V. vermiformis. Confocal microscopy demonstrated the intracellular presence of M. chimaera in V. vermiformis. CONCLUSION: In this way, V. vermiformis can contribute to the persistent presence of M. chimaera in HCUs. Cleaning and disinfection protocols should take this phenomenon into account.

Acanthamoeba castellanii interferes with adequate chlorine disinfection of multidrug-resistant Pseudomonas aeruginosa.

Verona-Integron-encoded-Metallo-ß-lactamase-positive Pseudomonas aeruginosa (VIM-PA) is a cause of hard-to-treat nosocomial infections, and can colonize hospital water networks alongside Acanthamoeba. We developed an in-vitro disinfection model to examine whether Acanthamoeba castellanii can harbour VIM-PA intracellularly, allowing VIM-PA to evade being killed by currently used hospital disinfectants. We observed that A. castellanii presence resulted in significantly increased survival of VIM-PA after exposure to chlorine for 30 s or for 2 min. This undesirable effect was not observed after disinfection by 70% alcohol or 24% acetic acid. Confocal microscopy confirmed the presence of VIM-PA within A. castellanii pseudocysts. Our data indicate that A. castellanii contributes to persistent VIM-PA colonization of water systems after chlorine treatment.

Haemostatic changes in urogenital schistosomiasis haematobium: a case-control study in Gabonese schoolchildren.

In many tropical areas schistosomiasis is a major health problem causing hepatosplenic, intestinal or urogenital complaints. Hepatosplenic schistosomiasis mansoni is also characterized by blood coagulation abnormalities. Liver pathology plays a role in the development of haemostatic changes and the parasitic infection may directly affect coagulation. However, these contributing factors cannot be studied separately in hepatosplenic schistosomiasis infections. This pilot study provides insight in haemostatic changes in urinary schistosomiasis by studying coagulation parameters in schistosomiasis haematobium-infected Gabonese schoolchildren. Selection on urinary schistosomiasis patients without hepatosplenic complaints allows for the investigation of the direct effects of the parasite on haemostasis. Levels of von Willebrand Factor (VWF) antigen, active VWF and osteoprotegerin were elevated, indicating inflammation-mediated endothelial activation. In contrast to hepatosplenic schistosomiasis, thrombin-antithrombin complex and D-dimer levels were not affected. Despite its small sample size, this study clearly indicates that Schistosoma haematobium directly alters the activation status of the endothelium, without initiation of coagulation.

Performance of the commercially available SERION ELISA classic Echinococcus IgG test for the detection of cystic echinococcosis in clinical practice.

Diagnosis of cystic echinococcosis (CE) is at present mainly based on imaging techniques. Serology has a complementary role, partly due to the small number of standardized and commercially available assays. Therefore we examined the clinical performance of the SERION ELISA classic Echinococcus IgG test. Using 10 U/ml as a cut-off point, and serum samples from 50 CE patients and 105 healthy controls, the sensitivity and specificity were 98.0% and 96.2%, respectively. If patients with other infectious diseases were used as negative controls, the specificity decreased to 76.9%, which causes poor positive predictive values. However, if results between 10 and 15 U/ml are classified as indecisive, the specificity of positive results (≥15 U/ml) increased to 92.5% without greatly affecting the sensitivity (92.0%). Using this approach in combination with imaging studies, the SERION ELISA classic Echinococcosis IgG test can be a useful aid in the diagnosis of CE.

Hypereosinophilia: a diagnostic challenge.

Hypereosinophilia encompasses a broad differential diagnosis of atopy/allergic reactions, drug reactions, parasitic infections and paraneoplastic syndromes. Although mostly of limited clinical significance, hypereosinophilia can also be related to hematological malignancies. One has to be aware of the potential for secondary organ damage for example, in the case of hypereosinophilic syndrome. We present three cases with different underlying mechanisms of hypereosinophilia with a brief overview of causes, diagnostic work-up and treatment options.

Recurrent amebic liver abscesses over a 16-year period: a case report.

BACKGROUND: Amebic liver abscess is a rare disease in high-income countries. Recurrence of amebic liver abscess is even rarer with only a few previous reports. Here we present a patient who developed three subsequent amebic liver abscesses over a sixteen-year period. CASE PRESENTATION: A Caucasian male developed recurrent amebic liver abscesses, when aged 23, 27 and 39 years. Only on the first occasion did this coincide with a recent visit to the tropics. The patient received adequate treatment during each episode. Possible explanations are persistent asymptomatic carrier state, cysts passage in his family, re-infection or chance. CONCLUSION: We describe the unusual case of a healthy male who developed recurrent amebic liver abscesses over a long period despite adequate treatment. Possible pathophysiological explanations are explored.

New insights in diagnosing Schistosoma myelopathy.

At present non-invasive tests for diagnosing Schistosoma myelopathy are sub-optimal. We present a novel serological method, using paired liquor and serum samples, resulting in the diagnosis of Schistosoma myelopathy in a male patient with proximal muscle weakness. The patient recovered after praziquantel treatment.

A review of photodynamic therapy in cutaneous leishmaniasis.

We present a review of six clinical studies investigating the use of photodynamic therapy (PDT) using porphyrin precursors for the treatment of Old World cutaneous leishmaniasis (CL). Thirty-nine patients with a total of 77 lesions received PDT using a range of treatment schedules following topical application of aminolevulinic acid (ALA) or methyl-aminolevulinate (MAL). The tissue response to PDT is accompanied by a mild burning sensation, erythema and reversible hypo- and hyperpigmentation. Few mechanistic studies have addressed the principles underlying the use of PDT for CL. All six reviewed papers suggest that PDT with porphyrin precursors is relatively effective in treating CL. Data are still limited, and PDT cannot at this point be recommended in routine clinical practice. The mechanism of action of this promising therapeutic modality needs to investigated further and additional controlled trials need to be performed.

Immunologic activity of schistosomal and bacterial TLR2 ligands in Gabonese children.

Schistosomes carry lipid moieties that interact with the immune system. To understand the consequence of interactions in terms of polarizing the cytokine profiles, the effect of two Toll-like receptor-2 (TLR2) activating schistosomal lipid fractions was studied on whole blood from Gabonese children living in a schistosomiasis endemic area. One fraction contained lysophosphatidylserine [monoacylglycerophosphoserine (lysoGPSer)] plus diacylphosphatidylserine [diacylglycerophosphoserine (GPSer)] while the other contained lysoGPSer and only a trace of GPSer. The effect of these schistosomal lipid fractions was compared with the known bacterial TLR2 ligands PAM3CSK4 and MALP-2. PAM3CSK4 and MALP-2 had preferential IL-10-activating capacities, while the fraction containing lysoGPSer plus GPSer had a strong TNF-alpha-inducing capacity. The fraction containing lysoGPSer was neutral with respect to pro- vs. anti-inflammatory effects. When Th1 and Th2 cytokines were analysed, the schistosomal lipid fraction containing lysoGPSer plus GPSer showed a stronger Th2 response compared to PAM3CSK4, MALP-2 and lysoGPSer alone. Therefore, the study indicates that not only TLR2 ligands derived from bacteria or from parasites can generate distinct cytokine profiles but also that the composition of lipid entities reaching the immune system can be important in leading to different immune outcomes. This information may be important for exploitation of immune modulatory molecules.

The extraordinary mitochondrion and unusual citric acid cycle in Trypanosoma brucei.

African trypanosomes are parasitic protozoa that cause sleeping sickness and nagana. Trypanosomes are not only of scientific interest because of their clinical importance, but also because these protozoa contain several very unusual biological features, such as their specially adapted mitochondrion and the compartmentalization of glycolytic enzymes in glycosomes. The energy metabolism of Trypanosoma brucei differs significantly from that of their hosts and changes drastically during the life cycle. Despite the presence of all citric acid cycle enzymes in procyclic insect-stage T. brucei, citric acid cycle activity is not used for energy generation. Recent investigations on the influence of substrate availability on the type of energy metabolism showed that absence of glycolytic substrates did not induce a shift from a fermentative metabolism to complete oxidation of substrates. Apparently, insect-stage T. brucei use parts of the citric acid cycle for other purposes than for complete degradation of mitochondrial substrates. Parts of the cycle are suggested to be used for (i) transport of acetyl-CoA units from the mitochondrion to the cytosol for the biosynthesis of fatty acids, (ii) degradation of proline and glutamate to succinate, (iii) generation of malate, which can then be used for gluconeogenesis. Therefore the citric acid cycle in trypanosomes does not function as a cycle.

Fasciola hepatica miracidia are dependent on respiration and endogenous glycogen degradation for their energy generation.

It is generally accepted that free-living stages of parasitic helminths are dependent on aerobic degradation of endogenous energy sources for their energy generation. This concept, however, is not the result of extensive experimental evidence, but originated mainly intuitively as oxygen is widely available in their habitat and these stages generally have a small size. Schistosoma mansoni, the sole parasitic helminth whose energy metabolism has been studied throughout its life-cycle indeed has aerobically functioning free-living stages. However, large differences exist in energy metabolism between adult stages of distinct parasitic helminths, and caution should be taken in predicting that all free-living stages of all parasitic helminths have the same, aerobic energy metabolism. Hence, this report studied the energy metabolism of Fasciola hepatica miracidia and demonstrated that F. hepatica miracidia are also dependent on aerobic degradation of their endogenous glycogen stores by glycolysis and on Krebs cycle activity for energy generation. However, in contrast to S. mansoni, F. hepatica miracidia cannot function anaerobically, as inhibition of the respiratory chain blocked motility and carbohydrate degradation, and finally resulted in death of the miracidia. Therefore, this report demonstrated that differences exist between miracidia of distinct species, in pre-adaptation of their energy metabolism to the occasional hypoxic conditions within their next host.

Isolation of Trypanosoma brucei CYC2 and CYC3 cyclin genes by rescue of a yeast G(1) cyclin mutant. Functional characterization of CYC2.

Two Trypanosoma brucei cyclin genes, CYC2 and CYC3, have been isolated by rescue of the Saccharomyces cerevisiae mutant DL1, which is deficient in CLN G(1) cyclin function. CYC2 encodes a 24-kDa protein that has sequence identity to the Neurospora crassa PREG1 and the S. cerevisiae PHO80 cyclin. CYC3 has the most sequence identity to mitotic B-type cyclins from a variety of organisms. Both CYC2 and CYC3 are single-copy genes and expressed in all life cycle stages of the parasite. To determine if CYC2 is found in a complex with previously identified trypanosome cdc2-related kinases (CRKs), the CYC2 gene was fused to the TY epitope tag, integrated into the trypanosome genome, and expressed under inducible control. CYC2ty was found to associate with an active trypanosome CRK complex since CYC2ty bound to leishmanial p12(cks1), and histone H1 kinase activity was detected in CYC2ty immune-precipitated fractions. Gene knockout experiments provide evidence that CYC2 is an essential gene, and co-immune precipitations together with a two-hybrid interaction assay demonstrated that CYC2 interacts with CRK3. The CRK3 x CYC2ty complex, the first cyclin-dependent kinase complex identified in trypanosomes, was localized by immune fluorescence to the cytoplasm throughout the cell cycle.

The CYC3 gene of trypanosoma brucei encodes a cyclin with a short half-life.

Recently, we identified two Trpanosoma brucei cyclin genes, CYC2 and CYC3, by rescue of the Saccharomyces cerevisiae mutant DL1, which is deficient in CLN G1 cyclin function. CYC3 has a low level of sequence identity to mitotic B-type cyclins from a variety of organisms. In order to examine whether CYC3 associates in vivo with a trypanosome cdc2-related kinase (CRK), the CYC3 gene was fused with the TY-epitope tag, integrated into the trypanosome genome and expressed under inducible control. CYC3ty was demonstrated to associate with the CRK-binding factor p12cks1 and histone H1 kinase activity could be detected in CYC3ty immune precipitated fractions, which demonstrates that CYC3ty associates in vivo with an active trypanosome CRK. Both CYC3ty and CYC2ty were shown to have a half-life of less than one cell cycle, which was significantly elongated by specific proteasome inhibitors, strongly suggesting that CYC3ty and CYC2ty are substrates for proteasome degradation. This is consistent with the presence in CYC3 of a putative destruction box motif that defines proteins for degradation via the ubiquitin degradation pathway. These results are consistant with proteolysis by the proteasome being involved in regulation of the cellular cyclin concentration in trypanosomes.

Ether lipids and their possible physiological function in adult Schistosoma mansoni.

Schistosomes have lost the capability to synthesize fatty acids de novo, but they can modify fatty acids by chain elongation. This has a profound effect on the molecular species composition of the two main phospholipid fractions of schistosomes, phosphatidylcholine (PC) and phosphatidylethanolamine (PE). Molecular species of phospholipids are increasingly recognized as important mediators, or precursors thereof, in signal transduction, immune response modulation, and events like membrane fusion. As these are all important aspects of schistosome membranes and of the tegumental membranes in particular, we analysed the PE and PC molecular species of the tegumental membranes, the worm body and the blood of the host. With the aid of on-line mass spectrometry, we unequivocally identified a large number of PC and PE species in schistosomes, among which considerable amounts of plasmalogen species. This was unexpected, as this lipid subclass has been assumed to be absent in the parasite. Species, like (20:1-16:0) diacyl PC and (16:0-20:1) plasmalogen PE, found to be main constituents in schistosomes, were absent from the blood of the host. Large differences were also found between the molecular species composition of the tegumental membranes and the membranes of the worm body. In the tegumental membranes, 1-hexadecyl 2-palmitoyl PC was detected, which could possibly function as a precursor for platelet activating factor (PAF).

A gene encoding the plant-like alternative oxidase is present in Phytomonas but absent in Leishmania spp.

The constituents of the respiratory chain are believed to differ among the trypanosomatids; bloodstream stages of African trypanosomes and Phytomonas promastigotes oxidize ubiquinol by a ubiquinol:oxygen oxidoreductase, also known as alternative oxidase, whereas Leishmania spp. oxidize ubiquinol via a classic cytochrome-containing respiratory chain. The molecular basis for this elementary difference in ubiquinol oxidation by the mitochondrial electron-transport chain in distinct trypanosomatids was investigated. The presence of a gene encoding the plant-like alternative oxidase could be demonstrated in Phytomonas and Trypanosoma brucei, trypanosomatids that are known to contain alternative oxidase activity. Our results further demonstrated that Leishmania spp. lack a gene encoding the plant-like alternative oxidase, and therefore, all stages of Leishmania spp. will lack the alternative oxidase protein. The observed fundamental differences between the respiratory chains of distinct members of the trypanosomatid family are thus caused by the presence or absence of a gene encoding the plant-like alternative oxidase.

The electron transport chain in anaerobically functioning eukaryotes.

Many lower eukaryotes can survive anaerobic conditions via a fermentation pathway that involves the use of the reduction of endogenously produced fumarate as electron sink. This fumarate reduction is linked to electron transport in an especially adapted, anaerobically functioning electron-transport chain. An aerobic energy metabolism with Krebs cycle activity is accompanied by electron transfer from succinate to ubiquinone via complex II of the respiratory chain. On the other hand, in an anaerobic metabolism, where fumarate functions as terminal electron acceptor, electrons are transferred from rhodoquinone to fumarate, which is the reversed direction. Ubiquinone cannot replace rhodoquinone in the process of fumarate reduction in vivo, as ubiquinone can only accept electrons from complex II and cannot donate them to fumarate. Rhodoquinone, with its lower redox potential than ubiquinone, is capable of donating electrons to fumarate. Eukaryotic fumarate reductases were shown to interact with rhodoquinone (a benzoquinone), whereas most prokaryotic fumarate reductases interact with the naphtoquinones menaquinone and demethylmenaquinone. Fumarate reductase, the enzyme essential for the anaerobic functioning of many eukaryotes, is structurally very similar to succinate dehydrogenase, the Krebs cycle enzyme catalysing the reverse reaction. In prokaryotes these enzymes are differentially expressed depending on the external conditions. Evidence is now emerging that also in eukaryotes two different enzymes exist for succinate oxidation and fumarate reduction that are differentially expressed.