MicrobiologyBytes

Trypanosomes are single-celled parasites that cause sleeping sickness in humans and wasting diseases in livestock. They are transmitted by the tsetse fly and, until now, it was unclear whether they reproduce sexually or asexually, because this stage in their life cycle occurs inside the insect carrier. Sexual reproduction produces offspring that inherit half their genetic material from each parent. The alternative is asexual reproduction, where the offspring inherit all genetic material from a single parent. Sexual reproduction is important in organisms that cause diseases because it can spread genes that make them more virulent, or resistant to drugs used for treatment, as well as creating completely new strains with combinations of genes not previously encountered. Some time ago it was shown that genetic shuffling could occur when two different trypanosome strains were mixed in the tsetse fly, but it was far from clear that this was true sexual reproduction. It was difficult to visualise the process directly because it happened inside the insect. To get round this problem, Professor Wendy Gibson and colleagues used fluorescently tagged proteins to make trypanosomes light up like tiny lightbulbs. They tagged proteins that function only during meiosis, the process of cellular division at the core of sexual reproduction that shuffles the parental genes and deals them out in new combinations to the offspring.

Read more: Caught in the act: trypanosome sex visualised for the first time

Identification of the meiotic life cycle stage of Trypanosoma brucei in the tsetse fly. PNAS USA February 14 2011 doi: 10.1073/pnas.1019423108

Wolbachia pipientis is an intracellular maternally-inherited bacterial symbiont of invertebrates that is very common in insects, including a number of mosquito species. It can manipulate host reproduction in several ways, including cytoplasmic incompatibility, whereby certain crosses are rendered effectively sterile. Females that are uninfected produce infertile eggs when they mate with males that carry Wolbachia, while there is a “rescue” effect in Wolbachia-infected embryos such that infected females can reproduce successfully with any males. Therefore uninfected females suffer a frequency-dependent reproductive disadvantage. Wolbachia is able to rapidly invade populations using this powerful mechanism

Malaria is one of the world’s most devastating diseases, particularly in Africa, and new control strategies are desperately needed. Here we show that the presence of Wolbachia bacteria inhibits the development of a malaria parasite in the most important Anopheles mosquito species of Africa. In addition it shows that the presence of Wolbachia results in the switching on of immune genes that are known to affect development of many species of malaria parasite. When added to the lifespan-shortening effects of this particular strain of Wolbachia, and the general ability of Wolbachia to spread through insect populations, this study provides a stimulus for the development of Wolbachia-based malaria control methods. It also provides new insights into the wide range of effects of Wolbachia in insects.

Wolbachia Stimulates Immune Gene Expression and Inhibits Plasmodium Development in Anopheles gambiae. (2010) PLoS Pathog 6(10): e1001143. doi:10.1371/journal.ppat.1001143
The over-replicating wMelPop strain of the endosymbiont Wolbachia pipientis has recently been shown to be capable of inducing immune upregulation and inhibition of pathogen transmission in Aedes aegypti mosquitoes. In order to examine whether comparable effects would be seen in the malaria vector Anopheles gambiae, transient somatic infections of wMelPop were created by intrathoracic inoculation. Upregulation of six selected immune genes was observed compared to controls, at least two of which (LRIM1 and TEP1) influence the development of malaria parasites. A stably infected An. gambiae cell line also showed increased expression of malaria-related immune genes. Highly significant reductions in Plasmodium infection intensity were observed in the wMelPop-infected cohort, and using gene knockdown, evidence for the role of TEP1 in this phenotype was obtained. Comparing the levels of upregulation in somatic and stably inherited wMelPop infections in Ae. aegypti revealed that levels of upregulation were lower in the somatic infections than in the stably transinfected line; inhibition of development of Brugia filarial nematodes was nevertheless observed in the somatic wMelPop infected females. Thus we consider that the effects observed in An. gambiae are also likely to be more pronounced if stably inherited wMelPop transinfections can be created, and that somatic infections of Wolbachia provide a useful model for examining effects on pathogen development or dissemination. The data are discussed with respect to the comparative effects on malaria vectorial capacity of life shortening and direct inhibition of Plasmodium development that can be produced by Wolbachia.

Related:

• Wolbachia
• Wolbachia could combat dengue fever
• Bacterial sequences in invertebrate genomes

Many organisms are able to adapt their development to the severity of their environment based on specific cues, and we have identified such a phenomenon, termed phenotypic plasticity, in the filarial parasite Litomosoides sigmodontis. Filarial nematodes infect about 200 million people worldwide, and much effort is going into finding a vaccine that would complement current drug treatments. Although anti-filarial immunity can be achieved, we show, in accord with evolutionary theory, that when these parasites infect a new host, they are able to adjust their development and reproduction to the presence of immune cells specialized in anti-helminth attack. These developmental schedules are determined within hours and impact their lifelong reproductive strategy; when immune attack is strong, and thus mortality is likely to be high, they produce offspring earlier and in greater numbers. Because current experimental vaccines rely on the very immune elements to which these nematodes adjust their development, their phenotypic plasticity could mitigate the expected reduction of disease burden in vaccinated populations.

Filarial Parasites Develop Faster and Reproduce Earlier in Response to Host Immune Effectors That Determine Filarial Life Expectancy. (2010) PLoS Biol 8(10): e1000525. doi:10.1371/journal.pbio.1000525
Humans and other mammals mount vigorous immune assaults against helminth parasites, yet there are intriguing reports that the immune response can enhance rather than impair parasite development. It has been hypothesized that helminths, like many free-living organisms, should optimize their development and reproduction in response to cues predicting future life expectancy. However, immune-dependant development by helminth parasites has so far eluded such evolutionary explanation. By manipulating various arms of the immune response of experimental hosts, we show that filarial nematodes, the parasites responsible for debilitating diseases in humans like river blindness and elephantiasis, accelerate their development in response to the IL-5 driven eosinophilia they encounter when infecting a host. Consequently they produce microfilariae, their transmission stages, earlier and in greater numbers. Eosinophilia is a primary host determinant of filarial life expectancy, operating both at larval and at late adult stages in anatomically and temporally separate locations, and is implicated in vaccine-mediated protection. Filarial nematodes are therefore able to adjust their reproductive schedules in response to an environmental predictor of their probability of survival, as proposed by evolutionary theory, thereby mitigating the effects of the immune attack to which helminths are most susceptible. Enhancing protective immunity against filarial nematodes, for example through vaccination, may be less effective at reducing transmission than would be expected and may, at worst, lead to increased transmission and, hence, pathology.

Related:

• Elimination of elephantiasis

Leishmania remains a major public health problem with 350 million people at risk, 12 million infected, and 2 million new infections per year. Despite the considerable progress in cellular and molecular biology and in evolutionary genetics since 1990, the debate on the population structure and reproductive mode of Leishmania is far from being settled and deserves further investigation. Two major hypotheses coexist: clonality versus sexuality. Because of the lack of clear evidence (experimental or biological confirmation) of sexuality in Leishmania parasites, until today it has been suggested and even accepted that Leishmania species were mainly clonal with infrequent genetic recombination.

Two recent publications, one on Leishmania major (an in vitro experimental study) and one on Leishmania braziliensis (a population genetics analysis), once again have challenged the hypothesis of clonal reproduction. The first study experimentally evidenced genetic recombination and proposed that Leishmania parasites are capable of having a sexual cycle consistent with meiotic processes inside the insect vector. The second investigation, based on population genetics studies, showed strong homozygosities, an observation that is incompatible with a predominantly clonal mode of reproduction at an ecological time scale (~20–500 generations). These studies highlight the need to advance the knowledge of Leishmania biology. This paper reviews the reasons stimulating the continued debate and then detail the next essential steps to be taken to clarify the Leishmania reproduction model. It widens the discussion to other Trypanosomatidae and show that the progress in Leishmania biology can improve our knowledge of the evolutionary genetics of American and African trypanosomes.

Everything You Always Wanted to Know about Sex (but Were Afraid to Ask) in Leishmania after Two Decades of Laboratory and Field Analyses. PLoS Pathog 6(8): e1001004. doi:10.1371/journal.ppat.1001004

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• Shedding light on the inner workings of the immune response
• Novel application for an old drug: tamoxifen in the treatment of Leishmania infection

Eukaryotic marine parasites could have a wide impact on marine ecology. This article in Microbiology Today describes the role of these parasites in processes as diverse as species competition, carbon cycling and gene transfer:

Parasites are typically small organisms that exploit their host both as a food source and as a habitat. Although well-studied as human pathogens and organisms prejudicial to human interests, they have been persistently ignored in microbial aquatic ecology. Increased awareness of the important role of viruses in marine aquatic ecosystems in processes as diverse as species competition, carbon cycling, and gene transfers has recently changed our overall view of aquatic parasites. Recent evidence of the widespread occurrence of small eukaryotic parasites, requiring eukaryotic hosts, has highlighted the existence of another kind of pathogen which potentially has specific ecological roles.

Read More

Related:

• Predator and Prey
• Microbial diseases and coral reefs

Plasmodium falciparum is a blood parasite that lives for the most part inside red cells. It is responsible for the death of 1-2 million people through malaria every year. The mechanisms by which the parasite invades red cells are complex and not completely understood. For many years it has been known that proteins called glycophorins are used by the parasite to gain entry into the red cell. However, the existence of another protein that allows entry independent of glycophorins has been suspected for nearly as long. The identity of the alternative protein has been a difficult mystery to solve.

Researchers have now identified an alternative protein used by P. falciparum to invade red blood cells. The results may aid the development of a future vaccine for malaria. The researchers identified complement receptor 1 as the protein that enables P. falciparum to invade red blood cells. Proteins called glycophorins are used by the parasite to gain entry into the red cell. However, because infection can take place without glycophorins, researchers suspected that another protein is also involved. Complement receptor 1 (CR1), also known to help protect red cells from attack by the immune system, has been suspected of having other roles in the development of malaria complications. The team was able to demonstrate that this protein is important in the invasion of red cells by using several laboratory strains of malaria as well as strains obtained from Kenya.

P. falciparum may use the CR1 protein instead of glycophorins if the parasite encounters a variant that lacks the glycophorin receptor; if the immune system mounts a response against parasite proteins involved in the dominant pathway due to a previous infection; or if the host were treated with a vaccine that blocks the glycophorin pathway. The recognition of the additional role of complement receptor 1 in red cell invasion will allow the definitive identification of malaria proteins that interact with it and that could be used in a future vaccine cocktail to block red cell invasion.

Complement Receptor 1 Is a Sialic Acid-Independent Erythrocyte Receptor of Plasmodium falciparum. 2010 PLoS Pathog 6(6): e1000968. doi:10.1371/journal.ppat.1000968
Plasmodium falciparum is a highly lethal malaria parasite of humans. A major portion of its life cycle is dedicated to invading and multiplying inside erythrocytes. The molecular mechanisms of erythrocyte invasion are incompletely understood. P. falciparum depends heavily on sialic acid present on glycophorins to invade erythrocytes. However, a significant proportion of laboratory and field isolates are also able to invade erythrocytes in a sialic acid-independent manner. The identity of the erythrocyte sialic acid-independent receptor has been a mystery for decades. We report here that the complement receptor 1 (CR1) is a sialic acid-independent receptor for the invasion of erythrocytes by P. falciparum. We show that soluble CR1 (sCR1) as well as polyclonal and monoclonal antibodies against CR1 inhibit sialic acid-independent invasion in a variety of laboratory strains and wild isolates, and that merozoites interact directly with CR1 on the erythrocyte surface and with sCR1- coated microspheres. Also, the invasion of neuraminidase-treated erythrocytes correlates with the level of CR1 expression. Finally, both sialic acid-independent and dependent strains invade CR1 transgenic mouse erythrocytes preferentially over wild-type erythrocytes but invasion by the latter is more sensitive to neuraminidase. These results suggest that both sialic acid-dependent and independent strains interact with CR1 in the normal red cell during the invasion process. However, only sialic acid-independent strains can do so without the presence of glycophorin sialic acid. Our results close a longstanding and important gap in the understanding of the mechanism of erythrocyte invasion by P. falciparum that will eventually make possible the development of an effective blood stage vaccine.

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• New ways to beat malaria
• Researchers characterize potential protein targets for malaria vaccine
• Beer consumption increases your attractiveness – to mosquitoes

The eukaryotic intestinal parasite Giardia intestinalis was first described in 1681, when Antonie van Leeuwenhoek undertook a microscopic examination of his own diarrhoeal stool. Nowadays, although G. intestinalis is recognized as a major worldwide contributor to diarrhoeal disease in humans and other mammals, the disease mechanisms are still poorly understood. Owing to its reduced complexity and proposed early evolutionary divergence, G. intestinalis is used as a model eukaryotic system for studying many basic cellular processes. This review discusses recent discoveries in the molecular cell biology and pathogenesis of G. intestinalis.

Behind the smile: cell biology and disease mechanisms of Giardia species. 2010 Nature Reviews Microbiology 8, 413-422 doi:10.1038/nrmicro2317

Related:

• Parasite genomes

Sleeping sickness, or Human African Trypanosomiasis, is a disease affecting the health and productivity of poor people in many rural areas of sub-Saharan Africa. The disease is caused by a single-celled flagellate, Trypanosoma brucei, which evades the immune system by periodically switching the proteins on its surface. Researchers have produced a genome sequence for T. brucei gambiense, which is the particular subspecies causing most disease in humans. They compared this with an existing reference genome for a non-human infecting strain to identify genes in T. b. gambiense that might explain its ability to infect humans and to assess how well the reference performs as a universal plan for all T. brucei. The genome sequences differ only due to rare insertions and duplications and homologous genes are over 95% identical on average. The archive of surface antigens that enable the parasite to switch its protein coat is remarkably consistent, even though it evolves very quickly. They also identified genes with predicted cell surface functions that are only present in T. b. brucei and have evolved rapidly in recent time. These genes might help to explain variation in disease pathology between different T. brucei strains in different hosts.

The team wanted to answer two questions: Is the existing T. b. brucei sequence representative of the full diversity of T. brucei parasites? And, is there anything in the T. b. gambiense genome that might explain its ability to infect and thrive in human populations? Historically, sleeping sickness has been a severely neglected disease, with considerable impact on human health and the well-being, and prosperity of communities. The genome comparison revealed a remarkable level of similarity between T. b. brucei and T. b. gambiense – just a single locus was unique to T. b. brucei. Moreover, the sequences of comparable genes were, on average, 98.2% identical. Because the genomes were so similar, the team could say with confidence that the T. b. brucei parasite and its genome are good models for future experiments to understand the biology of T. b. gambiense. The similarity between the two genomes also suggested that the source of T. b. gambiense’s ability to infect humans cannot be explained simply by the addition or removal of a few genes. Changes in the phenotype – the physical characteristics – seem to be down to more subtle changes in genetic information. Single letter changes in the genome; differences in the number of copies of genes; changes in how the activity of genes is regulated – all of these genetic nuances could play that crucial role in determining why T. b. gambiense behaves so differently to T. b. brucei. With two high-quality reference genome sequences in place for the T. brucei strains, the search for those small genetic differences is given a boost. It is this search that will fuel the pursuit of targeted drug treatments to tackle T. b. gambiense.

The Genome Sequence of Trypanosoma brucei gambiense, Causative Agent of Chronic Human African Trypanosomiasis. 2010 PLoS Negl Trop Dis 4(4): e658. doi:10.1371/journal.pntd.0000658

Related:

• Antigenic Variation in African Trypanosomes
• BILBO and the trypanosomes
• The Trypanosoma brucei flagellum

Leishmania donovani is a protozoan parasite that causes severe disease in humans with associated pathology in the spleen and liver. In experimental models of L. donovani infection, the hepatic response to infection is characterised by the presence of a focal mononuclear cell-rich inflammatory response (a granuloma) surrounding cells infected with intracellular amastigotes. Granulomas provide focus to the ensuing immune response, helping to contain parasite dissemination and providing the major effector site responsible for parasites elimination from the liver. Although granulomas are believed to form around infected resident liver macrophages (Kupffer cells), the role of these cells in intra-granuloma antigen presentation is currently unknown. Researchers used sophisticated microscopy to identify how killer T lymphocytes behaved when they enter sites of inflammation caused by L. donovani, and which infected cells they were able to recognise.

Leishmaniasis is a globally important but neglected disease, affecting approximately two million people every year. For most people, infection results in a slow-to-heal skin ulcer. In others, however, the parasite targets the liver, spleen and bone marrow, leading to over 70,000 deaths annually. The Leishmania parasite is eventually contained by a characteristic type of inflammatory response that forms cellular structures called granulomas. Little is known about the inner workings of these granulomas, in spite of their occurrence in other human diseases, from tuberculosis to rheumatoid arthritis. The scientists used an advanced laser-based microscopy technique, called “2-photon imaging”, to view the inner workings of the granuloma in mice infected with Leishmania. This enabled them to study how killer lymphocytes, such as those that can be induced by vaccination, are able to enter into the granulomas, penetrate deep into the core of the structure and seek out specific types of parasite-infected cells. Although this technique can not be used currently for the study of inflammatory disease in humans, the insights provided into the biology of granulomas and the hidden world of inflammation should help to improve vaccines and drugs, and allow researchers to now construct in silico models for this type of inflammatory process. These data have important implications for the understanding of how granulomas function to limit infection and may have important implications for the development of vaccines to Leishmania.

Dynamic Imaging of Experimental Leishmania donovani-Induced Hepatic Granulomas Detects Kupffer Cell-Restricted Antigen Presentation to Antigen-Specific CD8+ T Cells. PLoS Pathog 6(3): e1000805. doi:10.1371/journal.ppat.1000805
Kupffer cells (KCs) represent the major phagocytic population within the liver and provide an intracellular niche for the survival of a number of important human pathogens. Although KCs have been extensively studied in vitro, little is known of their in vivo response to infection and their capacity to directly interact with antigen-specific CD8+ T cells. Here, using a combination of approaches including whole mount and thin section confocal microscopy, adoptive cell transfer and intravital 2-photon microscopy, we demonstrate that KCs represent the only detectable population of mononuclear phagocytes within granulomas induced by Leishmania donovani infection that are capable of presenting parasite-derived peptide to effector CD8+ T cells. This restriction of antigen presentation to KCs within the Leishmania granuloma has important implications for the identification of new candidate vaccine antigens and for the design of novel immuno-therapeutic interventions.

Related:

• Novel application for an old drug: tamoxifen in the treatment of Leishmania infection
• Foamy macrophages allow TB agent to survive in infected individuals