Mauve Stingers (Pelagia noctiluca) as carriers of the bacterial fish pathogen Tenacibaculum maritimum

Aggregations or blooms of jellyfish are increasingly problematic for the aquaculture industry. Jellyfishassociated mass mortalities of sea-caged fish are most often caused by swarms of oceanic species like Pelagia noctiluca. These relatively large jellyfish get carried by tides and currents onto fish cages, causing them to break up into pathogenic nematocyst-containing pieces that are capable of passing through the mesh of the cages. The main effect on fish is gill damage leading to respiratory distress, but the lesions may also be compounded by bacterial infection, Tenacibaculum maritimum being one of the pathogens involved. In our previous study, we highlighted the ability of the jellyfish Phialella quadrata to carry this important pathogen. However, since these small jellyfish were collected around sea-cages of infected salmon, it was not possible to determine if the jellyfish or the fish themselves were the original source of the bacteria. Results of the current study demonstrate that these filamentous bacteria are present on the mouth of P. noctiluca that had no previous contact with farmed fish. These new results highlight the fact that some Cnidarian species harbour T. maritimum and suggest that jellyfishmight be a natural host for these bacteria whose environmental reservoir has not yet been determined.

CD18-dependent activation of the neutrophil NADPH oxidase during phagocytosis of Escherichia coli or Staphylococcus aureus is regulated by class III but not class I or II PI3Ks

Phagocytosis and activation of the NADPH oxidase are important mechanisms by which neutrophils and macrophages engulf and kill microbial pathogens. We investigated the role of PI3K signaling pathways in the regulation of the oxidase during phagocytosis of Staphylococcus aureus and Escherichia coli by mouse and human neutrophils, a mouse macrophage-like cell line and a human myeloid-like cell line. Phagocytosis of these bacteria was promoted by serum, independent of serum-derived antibodies, and effectively abolished in mouse neutrophils lacking the beta(2)-integrin common chain, CD18. A combination of PI3K isoform-selective inhibitors, mouse knock-outs, and RNA-interference indicated CD18-dependent activation of the oxidase was independent of class I and II PI3Ks, but substantially dependent on the single class III isoform (Vps34). Class III PI3K was responsible for the synthesis of PtdIns( 3) P on phagosomes containing either bacteria. The use of mouse neutrophils carrying an appropriate knock-in mutation indicated that PtdIns(3) P binding to the PX domain of their p40(phox) oxidase subunit is important for oxidase activation in response to both S aureus and E coli. This interaction does not, however, account for all the PI3K sensitivity of these responses, particularly the oxidase response to E coli, suggesting that additional mechanisms for PtdIns( 3) P-regulation of the oxidase must exist. ( Blood. 2008; 112: 5202-5211)

Development and clinical validation of multiplex TaqMan® assays for rapid diagnosis of viral gastroenteritis.

There is a need to provide rapid, sensitive, and often high throughput detection of pathogens in diagnostic virology. Viral gastroenteritis is a serious health issue often leading to hospitalization in the young, the immunocompromised and the elderly. The common causes of viral gastroenteritis include rotavirus, norovirus (genogroups I and II), astrovirus, and group F adenoviruses (serotypes 40 and 41). This article describes the work-up of two internally controlled multiplex, probe-based PCR assays and reports on the clinical validation over a 3-year period, March 2007 to February 2010. Multiplex assays were developed using a combination of TaqMan™ and minor groove binder (MGB™) hydrolysis probes. The assays were validated using a panel of 137 specimens, previously positive via a nested gel-based assay. The assays had improved sensitivity for adenovirus, rotavirus, and norovirus (97.3% vs. 86.1%, 100% vs. 87.8%, and 95.1% vs. 79.5%, respectively) and also more specific for targets adenovirus, rotavirus, and norovirus (99% vs. 95.2%, 100% vs. 93.6%, and 97.9% vs. 92.3%, respectively). For the specimens tested, both assays had equal sensitivity and specificity for astrovirus (100%). Overall the probe-based assays detected 16 more positive specimens than the nested gel-based assay. Post-introduction to the routine diagnostic service, a total of 9,846 specimens were processed with multiplex 1 and 2 (7,053 pediatric, 2,793 adult) over the 3-year study period. This clinically validated, probe-based multiplex testing algorithm allows highly sensitive and timely diagnosis of the four most prominent causes of viral gastroenteritis.

Genomic insights into the origin of parasitism in the emerging plant pathogen Bursaphelenchus xylophilus.

Bursaphelenchus xylophilus is the nematode responsible for a devastating epidemic of pine wilt disease in Asia and Europe, and represents a recent, independent origin of plant parasitism in nematodes, ecologically and taxonomically distinct from other nematodes for which genomic data is available. As well as being an important pathogen, the B. xylophilus genome thus provides a unique opportunity to study the evolution and mechanism of plant parasitism. Here, we present a high-quality draft genome sequence from an inbred line of B. xylophilus, and use this to investigate the biological basis of its complex ecology which combines fungal feeding, plant parasitic and insect-associated stages. We focus particularly on putative parasitism genes as well as those linked to other key biological processes and demonstrate that B. xylophilus is well endowed with RNA interference effectors, peptidergic neurotransmitters (including the first description of ins genes in a parasite) stress response and developmental genes and has a contracted set of chemosensory receptors. B. xylophilus has the largest number of digestive proteases known for any nematode and displays expanded families of lysosome pathway genes, ABC transporters and cytochrome P450 pathway genes. This expansion in digestive and detoxification proteins may reflect the unusual diversity in foods it exploits and environments it encounters during its life cycle. In addition, B. xylophilus possesses a unique complement of plant cell wall modifying proteins acquired by horizontal gene transfer, underscoring the impact of this process on the evolution of plant parasitism by nematodes. Together with the lack of proteins homologous to effectors from other plant parasitic nematodes, this confirms the distinctive molecular basis of plant parasitism in the Bursaphelenchus lineage. The genome sequence of B. xylophilus adds to the diversity of genomic data for nematodes, and will be an important resource in understanding the biology of this unusual parasite.

Rift Valley Fever Epidemiology, Surveillance, and Control: What Have Models Contributed?

Background: Rift Valley fever (RVF) is an emerging vector-borne zoonotic disease that represents a threat to human health, animal health, and livestock production, particularly in Africa. The epidemiology of RVF is not well understood, so that forecasting RVF outbreaks and carrying out efficient and timely control measures remains a challenge. Various epidemiological modeling tools have been used to increase knowledge on RVF epidemiology and to inform disease management policies.

Research strategies to improve honeybee health in Europe

Understanding the fundaments of colony losses and improving the status of colony health will require cross-cutting research initiatives including honeybee pathology, chemistry, genetics and apicultural extension. The 7th framework of the European Union requested research to empirically and experimentally fill knowledge gaps on honeybee pests and diseases, including 'Colony Collapse Disorder' and the impact of parasites, pathogens and pesticides on honeybee mortality. The interactions among these drivers of colony loss will be studied in different European regions, using experimental model systems including selected parasites (e. g. Nosema and Varroa mites), viruses (Deformed Wing Virus, Black Queen Cell Virus, Israeli Acute Paralysis Virus) and model pesticides (thiacloprid, tau-fluvalinate). Transcriptome analyses will be used to explore host-pathogen-pesticide interactions and identify novel genes for disease resistance. Special attention will be given to sublethal and chronic exposure to pesticides and will screen how apicultural practices affect colony health. Novel diagnostic screening methods and sustainable concepts for disease prevention will be developed resulting in new treatments and selection tools for resistant stock. Research initiatives will be linked to various national and international ongoing European, North-and South-American colony health monitoring and research programs, to ensure a global transfer of results to apicultural practice in the world community of beekeepers.

The conservation of bees: a global perspective

Bees are major pollinators of Angiosperms and therefore their apparent decline is of importance for humans and biodiversity. We synthesise results of 12 recent reviews to provide a global picture of the threats they face. Habitat loss is the major threat to bee diversity, whilst invasive species, emerging diseases, pesticide use, and climate change also have the potential to impact bee populations. We suggest that future conservation strategies need to prioritise (i) minimising habitat loss, (ii) making agricultural habitats bee-friendly, (iii) training scientists and the public in bee taxonomy and identification, (iv) basic autecological and population genetic studies to underpin conservation strategies, (v) assessing the value of DNA barcoding for bee conservation, (vi) determining the impact of invasive plants, animals, parasites and pathogens, and (vii) integrating this information to understand the potential impact of climate change on current bee diversity.

Early target cells of measles virus after aerosol infection of non-human primates

Measles virus (MV) is highly infectious, and has long been thought to enter the host by infecting epithelial cells of the respiratory tract. However, epithelial cells do not express signaling lymphocyte activation molecule (CD150), which is the high-affinity cellular receptor for wild-type MV strains. We have generated a new recombinant MV strain expressing enhanced green fluorescent protein (EGFP), based on a wild-type genotype B3 virus isolate from Khartoum, Sudan (KS). Cynomolgus macaques were infected with a high dose of rMV(KS)EGFP by aerosol inhalation to ensure that the virus could reach the full range of potential target cells throughout the entire respiratory tract. Animals were euthanized 2, 3, 4 or 5 days post-infection (d.p.i., n?=?3 per time point) and infected (EGFP(+)) cells were identified at all four time points, albeit at low levels 2 and 3 d.p.i. At these earliest time points, MV-infected cells were exclusively detected in the lungs by fluorescence microscopy, histopathology and/or virus isolation from broncho-alveolar lavage cells. On 2 d.p.i., EGFP(+) cells were phenotypically typed as large mononuclear cells present in the alveolar lumen or lining the alveolar epithelium. One to two days later, larger clusters of MV-infected cells were detected in bronchus-associated lymphoid tissue (BALT) and in the tracheo-bronchial lymph nodes. From 4 d.p.i. onward, MV-infected cells were detected in peripheral blood and various lymphoid tissues. In spite of the possibility for the aerosolized virus to infect cells and lymphoid tissues of the upper respiratory tract, MV-infected cells were not detected in either the tonsils or the adenoids until after onset of viremia. These data strongly suggest that in our model MV entered the host at the alveolar level by infecting macrophages or dendritic cells, which traffic the virus to BALT or regional lymph nodes, resulting in local amplification and subsequent systemic dissemination by viremia.

Helminth Cysteine Proteases Inhibit TRIF-dependent Activation of Macrophages via Degradation of TLR3

Helminth pathogens prepare a Th2 type immunological environment in their hosts to ensure their longevity. They achieve this by secreting molecules that not only actively drive type 2 responses but also suppress type 1 responses. Here, we show that the major cysteine proteases secreted from the helminth pathogens Fasciola hepatica (FheCL1) and Schistosoma mansoni (SmCB1) protect mice from the lethal effects of lipopolysaccharide by preventing the release of inflammatory mediators, nitric oxide, interleukin-6, tumor necrosis factor alpha, and interleukin-12, from macrophages. The proteases specifically block the MyD88-independent TRIF-dependent signaling pathway of Toll-like receptor (TLR) 4 and TLR3. Microscopical and flow cytometric studies, however, show that alteration of macrophage function by cysteine protease is not mediated by cleavage of components of the TLR4 complex on the cell surface but occurs by degradation of TLR3 within the endosome. This is the first study to describe a parasite molecule that degrades this receptor and pinpoints a novel mechanism by which helminth parasites modulate the innate immune responses of their hosts to suppress the development of Th1 responses.

A Family of Helminth Molecules that Modulate Innate Cell Responses via Molecular Mimicry of Host Antimicrobial Peptides

Over the last decade a significant number of studies have highlighted the central role of host antimicrobial (or defence) peptides in modulating the response of innate immune cells to pathogen-associated ligands. In humans, the most widely studied antimicrobial peptide is LL-37, a 37-residue peptide containing an amphipathic helix that is released via proteolytic cleavage of the precursor protein CAP18. Owing to its ability to protect against lethal endotoxaemia and clinically-relevant bacterial infections, LL-37 and its derivatives are seen as attractive candidates for anti-sepsis therapies. We have identified a novel family of molecules secreted by parasitic helminths (helminth defence molecules; HDMs) that exhibit similar biochemical and functional characteristics to human defence peptides, particularly CAP18. The HDM secreted by Fasciola hepatica (FhHDM-1) adopts a predominantly alpha-helical structure in solution. Processing of FhHDM-1 by F. hepatica cathepsin L1 releases a 34-residue C-terminal fragment containing a conserved amphipathic helix. This is analogous to the proteolytic processing of CAP18 to release LL-37, which modulates innate cell activation by classical toll-like receptor (TLR) ligands such as lipopolysaccharide (LPS). We show that full-length recombinant FhHDM-1 and a peptide analogue of the amphipathic C-terminus bind directly to LPS in a concentration-dependent manner, reducing its interaction with both LPS-binding protein (LBP) and the surface of macrophages. Furthermore, FhHDM-1 and the amphipathic C-terminal peptide protect mice against LPS-induced inflammation by significantly reducing the release of inflammatory mediators from macrophages. We propose that HDMs, by mimicking the function of host defence peptides, represent a novel family of innate cell modulators with therapeutic potential in anti-sepsis treatments and prevention of inflammation.

The repertoire of G protein-coupled receptors in the human parasite Schistosoma mansoni and the model organism Schmidtea mediterranea.

Background

G protein-coupled receptors (GPCRs) constitute one of the largest groupings of eukaryotic proteins, and represent a particularly lucrative set of pharmaceutical targets. They play an important role in eukaryotic signal transduction and physiology, mediating cellular responses to a diverse range of extracellular stimuli. The phylum Platyhelminthes is of considerable medical and biological importance, housing major pathogens as well as established model organisms. The recent availability of genomic data for the human blood fluke Schistosoma mansoni and the model planarian Schmidtea mediterranea paves the way for the first comprehensive effort to identify and analyze GPCRs in this important phylum.

Results

Application of a novel transmembrane-oriented approach to receptor mining led to the discovery of 117 S. mansoni GPCRs, representing all of the major families; 105 Rhodopsin, 2 Glutamate, 3 Adhesion, 2 Secretin and 5 Frizzled. Similarly, 418 Rhodopsin, 9 Glutamate, 21 Adhesion, 1 Secretin and 11 Frizzled S. mediterranea receptors were identified. Among these, we report the identification of novel receptor groupings, including a large and highly-diverged Platyhelminth-specific Rhodopsin subfamily, a planarian-specific Adhesion-like family, and atypical Glutamate-like receptors. Phylogenetic analysis was carried out following extensive gene curation. Support vector machines (SVMs) were trained and used for ligand-based classification of full-length Rhodopsin GPCRs, complementing phylogenetic and homology-based classification.

Conclusions

Genome-wide investigation of GPCRs in two platyhelminth genomes reveals an extensive and complex receptor signaling repertoire with many unique features. This work provides important sequence and functional leads for understanding basic flatworm receptor biology, and sheds light on a lucrative set of anthelmintic drug targets.

Reducing the future threat from (liver) fluke: realistic prospect or quixotic fantasy?

The liver fluke remains an economically significant parasite of livestock and is emerging as an important zoonotic infection of humans. The incidence of the disease has increased in the last few years, as a possible consequence of changes to the World's climate. Future predictions suggest that this trend is likely to continue. Allied to the changing pattern of disease, reports of resistance to triclabendazole (TCBZ) have appeared in the literature, although they do not all represent genuine cases of resistance. Nevertheless, any reports of resistance are a concern, because triclabendazole is the only drug that has high activity against the migratory and damaging juvenile stages of infection. How to deal with the twin problems (of increasing incidence and drug resistance) is the overall theme of the session on “Trematodes: Fasciola hepatica epidemiology and control” and of this review to introduce the session.

Greater knowledge of fluke epidemiology and population genetics will highlight those regions where surveillance is most required and indicate how quickly resistant populations of fluke may arise. Models of disease risk are becoming increasingly sophisticated and precise, with more refined data analysis programmes and Geographic Information Systems (GIS) data. Recent improvements have been made in our understanding of the action of triclabendazole and the ways in which flukes have become resistant to it. While microtubules are the most likely target for drug action, tubulin mutations do not seem to be involved in the resistance mechanism. Rather, upregulation of drug uptake and metabolism processes appear to be more important and the data relating to them will be discussed. The information may help in the design of new treatment strategies or pinpoint potential molecular markers for monitoring fluke populations. Advances in the identification of novel targets for drugs and vaccines will be made by the various “-omics” technologies that are now being applied to Fasciola. A major area of concern in the current control of fasciolosis is the lack of reliable tests for the diagnosis of drug (TCBZ) resistance. This has led to inaccurate reports of resistance, which is hindering successful disease management, as farmers may be encouraged to switch to less effective drugs. Progress with the development of a number of new diagnostic tests will be reviewed.

Treatment of estrogens and androgens in dairy wastewater by a constructed wetland system.

Constructed wetland systems (CWS) have been used as a low cost bio-filtration system to treat farm wastewater. While studies have shown that CWS are efficient in removing organic compounds and pathogens, there is limited data on the presence of hormones in this type of treatment system. The objective of this study was to evaluate the ability of the CWS to reduce estrogenic and androgenic hormone concentration in dairy wastewater. This was achieved through a year long study on dairy wastewater samples obtained froma surface flow CWS. Analysis of hormonal levels was performed using a solid phase extraction (SPE) sample clean-up method, combined with reporter gene assays (RGAs) which incorporate relevant receptors capable of measuring total estrogenic or androgenic concentrations as low as 0.24 ng L1 and 6.9 ng L1 respectively. Monthly analysis showed a mean removal efficiency for estrogens of 95.2%, corresponding to an average residual concentration of 3.2 ng L1 17b-estradiol equivalent (EEQ), below the proposed lowest observable effect concentration (LOEC) of 10 ng L1. However, for one month a peak EEQ concentration of 115 ng L1 was only reduced to 18.8 ng L1. The mean androgenic activity peaked at 360 ng L1 and a removal efficiency of 92.1% left an average residual concentration of 32.3 ng L1 testosterone equivalent (TEQ). The results obtained demonstrate that this type of CWS is an efficient system for the treatment of hormones in dairy wastewater. However, additional design improvements may be required to further enhance removal efficiency of peak hormone concentrations.