Differences between <i>Trypanosoma brucei gambiense</i> groups 1 and 2 in their resistance to killing by Trypanolytic factor 1

&lt;p&gt;&lt;b&gt;Background:&lt;/b&gt; The three sub-species of &lt;i&gt;Trypanosoma brucei&lt;/i&gt; are important pathogens of sub-Saharan Africa. &lt;i&gt;T. b. brucei&lt;/i&gt; is unable to infect humans due to sensitivity to trypanosome lytic factors (TLF) 1 and 2 found in human serum. &lt;i&gt;T. b. rhodesiense&lt;/i&gt; and &lt;i&gt;T. b. gambiense&lt;/i&gt; are able to resist lysis by TLF. There are two distinct sub-groups of &lt;i&gt;T. b. gambiense&lt;/i&gt; that differ genetically and by human serum resistance phenotypes. Group 1 &lt;i&gt;T. b. gambiense&lt;/i&gt; have an invariant phenotype whereas group 2 show variable resistance. Previous data indicated that group 1 &lt;i&gt;T. b. gambiense&lt;/i&gt; are resistant to TLF-1 due in-part to reduced uptake of TLF-1 mediated by reduced expression of the TLF-1 receptor (the haptoglobin-hemoglobin receptor (&lt;i&gt;HpHbR&lt;/i&gt;)) gene. Here we investigate if this is also true in group 2 parasites.&lt;/p&gt; &lt;p&gt;&lt;b&gt;Methodology:&lt;/b&gt; Isogenic resistant and sensitive group 2 &lt;i&gt;T. b. gambiense&lt;/i&gt; were derived and compared to other T. brucei parasites. Both resistant and sensitive lines express the &lt;i&gt;HpHbR&lt;/i&gt; gene at similar levels and internalized fluorescently labeled TLF-1 similar fashion to &lt;i&gt;T. b. brucei&lt;/i&gt;. Both resistant and sensitive group 2, as well as group 1 &lt;i&gt;T. b. gambiense&lt;/i&gt;, internalize recombinant APOL1, but only sensitive group 2 parasites are lysed.&lt;/p&gt; &lt;p&gt;&lt;b&gt;Conclusions:&lt;/b&gt; Our data indicate that, despite group 1 &lt;i&gt;T. b. gambiense&lt;/i&gt; avoiding TLF-1, it is resistant to the main lytic component, APOL1. Similarly group 2 &lt;i&gt;T. b. gambiense&lt;/i&gt; is innately resistant to APOL1, which could be based on the same mechanism. However, group 2 &lt;i&gt;T. b. gambiense&lt;/i&gt; variably displays this phenotype and expression does not appear to correlate with a change in expression site or expression of &lt;i&gt;HpHbR&lt;/i&gt;. Thus there are differences in the mechanism of human serum resistance between &lt;i&gt;T. b. gambiense&lt;/i&gt; groups 1 and 2.&lt;/p&gt

Trypanosoma brucei gambiense group 1 is distinguished by a unique amino acid substitution in the HpHb receptor implicated in human serum resistance

Trypanosoma brucei rhodesiense (Tbr) and T. b. gambiense (Tbg), causative agents of Human African Trypanosomiasis (sleeping sickness) in Africa, have evolved alternative mechanisms of resisting the activity of trypanosome lytic factors (TLFs), components of innate immunity in human serum that protect against infection by other African trypanosomes. In Tbr, lytic activity is suppressed by the Tbr-specific serum-resistance associated (SRA) protein. The mechanism in Tbg is less well understood but has been hypothesized to involve altered activity and expression of haptoglobin haemoglobin receptor (HpHbR). HpHbR has been shown to facilitate internalization of TLF-1 in T.b. brucei (Tbb), a member of the T. brucei species complex that is susceptible to human serum. By evaluating the genetic variability of HpHbR in a comprehensive geographical and taxonomic context, we show that a single substitution that replaces leucine with serine at position 210 is conserved in the most widespread form of Tbg (Tbg group 1) and not found in related taxa, which are either human serum susceptible (Tbb) or known to resist lysis via an alternative mechanism (Tbr and Tbg group 2). We hypothesize that this single substitution contributes to reduced uptake of TLF and thus may play a key role in conferring serum resistance to Tbg group 1. In contrast, similarity in HpHbR sequence among isolates of Tbg group 2 and Tbb/Tbr provides further evidence that human serum resistance in Tbg group 2 is likely independent of HpHbR functio

Program Model Checking: A Practitioner's Guide

Program model checking is a verification technology that uses state-space exploration to evaluate large numbers of potential program executions. Program model checking provides improved coverage over testing by systematically evaluating all possible test inputs and all possible interleavings of threads in a multithreaded system. Model-checking algorithms use several classes of optimizations to reduce the time and memory requirements for analysis, as well as heuristics for meaningful analysis of partial areas of the state space Our goal in this guidebook is to assemble, distill, and demonstrate emerging best practices for applying program model checking. We offer it as a starting point and introduction for those who want to apply model checking to software verification and validation. The guidebook will not discuss any specific tool in great detail, but we provide references for specific tools

Gene Expression Profiles of Chlamydophila pneumoniae during the Developmental Cycle and Iron Depletion–Mediated Persistence

The obligate intracellular, gram-negative bacterium Chlamydophila pneumoniae (Cpn) has impact as a human pathogen. Little is known about changes in the Cpn transcriptome during its biphasic developmental cycle (the acute infection) and persistence. The latter stage has been linked to chronic diseases. To analyze Cpn CWL029 gene expression, we designed a pathogen-specific oligo microarray and optimized the extraction method for pathogen RNA. Throughout the acute infection, ratio expression profiles for each gene were generated using 48 h post infection as a reference. Based on these profiles, significantly expressed genes were separated into 12 expression clusters using self-organizing map clustering and manual sorting into the “early”, “mid”, “late”, and “tardy” cluster classes. The latter two were differentiated because the “tardy” class showed steadily increasing expression at the end of the cycle. The transcriptome of the Cpn elementary body (EB) and published EB proteomics data were compared to the cluster profile of the acute infection. We found an intriguing association between “late” genes and genes coding for EB proteins, whereas “tardy” genes were mainly associated with genes coding for EB mRNA. It has been published that iron depletion leads to Cpn persistence. We compared the gene expression profiles during iron depletion–mediated persistence with the expression clusters of the acute infection. This led to the finding that establishment of iron depletion–mediated persistence is more likely a mid-cycle arrest in development rather than a completely distinct gene expression pattern. Here, we describe the Cpn transcriptome during the acute infection, differentiating “late” genes, which correlate to EB proteins, and “tardy” genes, which lead to EB mRNA. Expression profiles during iron mediated–persistence led us to propose the hypothesis that the transcriptomic “clock” is arrested during acute mid-cycle

Detection of Schmallenberg virus in Culicoides species of the Obsoletus complex in Germany

Malformed lambs were born on an animal husbandry station in Brandenburg, Germany in January 2012 with signs typical of a Schmallenberg virus (SBV) infection such as torticollis, arthrogryposis and inferior brachygnathia. Infections with SBV were confirmed by post-mortem and laboratory diagnosis. This finding prompted us to retrospectively investigate the eventual presence of SBV in insects collected on pastures of the farm during 2011 while conducting two entomological studies. Insects were collected at dusk with a drop trap from cattle in the first study (16 May - 8 July 2011) and from sheep in the second study (19 September - 5 October 2011) and stored in 70% ethanol. Genome sequences of SBV were detected by a real time quantitative reverse transcription PCR (RT-qPCR) in a pool of biting midges belonging to the Culicoides obsoletus complex that were caught in the week of 26 September 2011. All pools of Aedes and Culex mosquitoes and the Culicoides spp. collected during the first study in May-July 2011 tested negative

Managing Tsetse Transmitted Trypanosomosis by Insecticide Treated Nets - an Affordable and Sustainable Method for Resource Poor Pig Farmers in Ghana

An outbreak of tsetse-transmitted trypanosomiasis resulted in more than 50% losses of domestic pigs in the Eastern Region of Ghana (source: Veterinary Services, Accra; April 2007). In a control trial from May 4th–October 10th 2007, the efficacy of insecticide-treated mosquito fences to control tsetse was assessed. Two villages were selected – one serving as control with 14 pigsties and one experimental village where 24 pigsties were protected with insecticide treated mosquito fences. The 100 cm high, 150denier polyester fences with 100 mg/m2 deltamethrin and a UV protector were attached to surrounding timber poles and planks. Bi-monthly monitoring of tsetse densities with 10 geo-referenced bi-conical traps per village showed a reduction of more than 90% in the protected village within two months. Further reductions exceeding 95% were recorded during subsequent months. The tsetse population in the control village was not affected, only displaying seasonal variations. Fifty pigs from each village were ear-tagged and given a single curative treatment with diminazene aceturate (3.5 mg/kg bw) after their blood samples had been taken. The initial trypanosome prevalence amounted to 76% and 72% of protected and control animals, respectively, and decreased to 16% in protected as opposed to 84% in control pigs three months after intervention. After six months 8% of the protected pigs were infected contrasting with 60% in the control group

Phylogeography and Taxonomy of Trypanosoma brucei

Trypanosoma brucei, the parasite causing human African trypanosomiasis (sleeping sickness) across sub-Saharan Africa is traditionally split into three subspecies: T. b. gambiense (Tbg), causing a chronic form of human disease in West and Central Africa; T. b. rhodesiense (Tbr), causing an acute form of human disease in East and Southern Africa; and T. b. brucei (Tbb), which is restricted to animals. Tbg is further split into Tbg group 1 and Tbg group 2. Better understanding the evolutionary relationships between these groups may help to shed light on the epidemiology of sleeping sickness. Here, we used three different types of genetic markers to investigate the phylogeographic relationships among the four groups across a large portion of their range. Our results confirm the distinctiveness of Tbg group 1 while highlighting the extremely close relationships among the other three taxa. In particular, Tbg group 2 was closely related to Tbb, while Tbr appeared to be a variant of Tbb, differing only in its phenotype of human infectivity. The wide geographic distribution of the gene conferring human infectivity (SRA) and the fact that it is readily exchanged among lineages of T. brucei in eastern Africa suggests that human-infective trypanosomes have access to an extensive gene pool with which to respond to selective pressures such as drugs

The Animal Trypanosomiases and their chemotherapy:a review

Pathogenic animal trypanosomes affecting livestock have represented a major constraint to agricultural development in Africa for centuries, and their negative economic impact is increasing in South America and Asia. Chemotherapy and chemoprophylaxis represent the main means of control. However, research into new trypanocides has remained inadequate for decades, leading to a situation where the few compounds available are losing efficacy due to the emergence of drug-resistant parasites. In this review, we provide a comprehensive overview of the current options available for the treatment and prophylaxis of the animal trypanosomiases, with a special focus on the problem of resistance. The key issues surrounding the main economically important animal trypanosome species and the diseases they cause are also presented. As new investment becomes available to develop improved tools to control the animal trypanosomiases, we stress that efforts should be directed towards a better understanding of the biology of the relevant parasite species and strains, to identify new drug targets and interrogate resistance mechanisms