The role of TLR2 in cutaneous leishmaniasis and as a target for vaccine adjuvants

After the introduction of clean water, vaccination is thought to be the most effective public health tool ever introduced, responsible for preventing millions of cases of disease, disability and death each year. Unfortunately there remain a number of important human diseases for which we have no vaccine, particularly parasitic diseases, such as leishmaniasis, which primarily affect poor communities in tropical regions. There are many complex reasons why we have failed to develop effective vaccines for parasitic diseases, but there is hope that with our improved understanding of the immune system alongside the development of a new generation of vaccines, we will soon develop new vaccines which are effective enough to prevent such diseases. Toll-like receptors (TLRs) are major targets for adjuvants and have been shown to be crucial for defence against a number of infections. TLR2 recognises bacterial lipopeptides in a heterodimer with either TLR1 or TLR6, and its function has been linked to protection against various bacterial infections and to the efficacy of the BCG vaccine. TLR2 has been shown to recognise surface glycoconjugates of Leishmania parasites in vitro, particularly lipophosphoglycan (LPG). In this study, in vivo experimental infections show that TLR2 has a protective role in controlling cutaneous leishmaniasis (CL), as shown by increased lesion sizes and parasite burdens in TLR2-/- mice infected with L. major and L. mexicana. Furthermore, it appears that LPG is not the major mediator of TLR2 activation during infection with L. mexicana, as parasites lacking LPG also resulted in exacerbated disease in TLR2-/- mice. Mice lacking TLR2 co-receptors TLR1 and TLR6 did not show increased susceptibility to infection, suggesting either mono-TLR2 function or alternative co-receptor involvement. Infected TLR2-/- mice show a skewed Th2 immune response to Leishmania, as demonstrated by elevated IL-4, IL-13 and IL-10 production by draining lymph node (DLN) cells in response to antigen. These results suggest that TLR2 is involved in promoting protective immune responses to Leishmania parasites during primary infection in vivo, and is a potential target for protective and therapeutic vaccine adjuvants. Paradoxically, however, TLR2-targeting lipopeptides Pam2 and Pam3 were ineffective adjuvants for use in a whole-cell vaccine to protect against CL, as whole-cell autoclaved L. major (ALM) vaccines containing lipopeptides resulted in exacerbated disease upon challenge when compared to unvaccinated controls and in contrast to effective vaccination when CpG adjuvants were used. The ratio of antigen specific IgG1:IgG2c antibody isotypes, which is a marker of the type of adaptive immune response (Th1 or Th2), was elevated in mice that received vaccines containing lipopeptide adjuvants, suggesting that these adjuvants drive non-protective Th2 responses to Leishmania. In a Th2-dependent vaccine model using Brugia malayi, the use of Pam2 as an adjuvant resulted in an enhanced protective phenotype with similar efficacy to the Th2-driving adjuvant Alum. Thus, in the context of CL infection TLR2 has a protective role in late-stage primary infections with L. major and L. mexicana, yet when targeted with lipopeptide adjuvants in whole-cell vaccines promotes exacerbated disease in challenge infections, through driving Th2 immune responses. Lipopeptides that target TLR2, such as Pam2, are therefore more appropriate for use as adjuvants in vaccines where Th2 protective immunity is required

Predicting progression to active tuberculosis:A rate-limiting step on the path to elimination

In a Perspective, Ajit Lalvani and colleagues discuss new approaches to predicting progression to active tuberculosis

Eosinophil-Mediated Immune Control of Adult Filarial Nematode Infection Can Proceed in the Absence of IL-4 Receptor Signaling

Helminth infections are accompanied by eosinophilia in parasitized tissues. Eosinophils are effectors of immunity to tissue helminths. We previously reported that in the context of experimental filarial nematode infection, optimum tissue eosinophil recruitment was coordinated by local macrophage populations following IL-4R–dependent in situ proliferation and alternative activation. However, in the current study, we identify that control of chronic adult filarial worm infection is evident in IL-4Ra–deficient (IL-4Ra2/2) mice, whereby the majority of infections do not achieve patency. An associated residual eosinophilia was apparent in infected IL-4Ra2/2 mice. By treating IL-4Ra2/2 mice serially with anti-CCR3 Ab or introducing a compound deficiency in CCR3 within IL-4Ra2/2 mice, residual eosinophilia was ablated, and susceptibility to chronic adult Brugia malayi infection was established, promoting a functional role for CCR3-dependent eosinophil influx in immune control in the absence of IL-4/IL13–dependent immune mechanisms. We investigated additional cytokine signals involved in residual eosinophilia in the absence IL-4Ra signaling and defined that IL-4Ra2/2/IL-52/2 double-knockout mice displayed significant eosinophil deficiency compared with IL-4Ra2/2 mice and were susceptible to chronic fecund adult filarial infections. Contrastingly, there was no evidence that either IL-4R–dependent or IL-4R–independent/CCR3/IL-5–dependent immunity influenced B. malayi microfilarial loads in the blood. Our data demonstrate multiplicity of Th2-cytokine control of eosinophil tissue recruitment during chronic filarial infection and that IL-4R–independent/IL-5– and CCR3-dependent pathways are sufficient to control filarial adult infection via an eosinophil-dependent effector response prior to patency

Interleukin-4 activated macrophages mediate immunity to filarial helminth infection by sustaining CCR3-dependent eosinophilia

Eosinophils are effectors in immunity to tissue helminths but also induce allergic immunopathology. Mechanisms of eosinophilia in non-mucosal tissues during infection remain unresolved. Here we identify a pivotal function of tissue macrophages (Mϕ) in eosinophil anti-helminth immunity using a BALB/c mouse intra-peritoneal Brugia malayi filarial infection model. Eosinophilia, via C-C motif chemokine receptor (CCR)3, was necessary for immunity as CCR3 and eosinophil impairments rendered mice susceptible to chronic filarial infection. Post-infection, peritoneal Mϕ populations proliferated and became alternatively-activated (AAMϕ). Filarial AAMϕ development required adaptive immunity and interleukin-4 receptor-alpha. Depletion of Mϕ prior to infection suppressed eosinophilia and facilitated worm survival. Add back of filarial AAMϕ in Mϕ-depleted mice recapitulated a vigorous eosinophilia. Transfer of filarial AAMϕ into Severe-Combined Immune Deficient mice mediated immunological resistance in an eosinophil-dependent manner. Exogenous IL-4 delivery recapitulated tissue AAMϕ expansions, sustained eosinophilia and mediated immunological resistance in Mϕ-intact SCID mice. Co-culturing Brugia with filarial AAMϕ and/or filarial-recruited eosinophils confirmed eosinophils as the larvicidal cell type. Our data demonstrates that IL-4/IL-4Rα activated AAMϕ orchestrate eosinophil immunity to filarial tissue helminth infection

A Study of Interstellar Gas and Stars in the Gravitationally Lensed Galaxy `The Cosmic Eye' from Rest-Frame Ultraviolet Spectroscopy

We report the results of a study of the rest-frame UV spectrum of the Cosmic Eye, a luminous Lyman break galaxy at z=3.07331 gravitationally lensed by a factor of 25. The spectrum, recorded with the ESI spectrograph on the Keck II telescope, is rich in absorption features from the gas and massive stars in this galaxy. The interstellar absorption lines are resolved into two components of approximately equal strength and each spanning several hundred km/s in velocity. One component has a net blueshift of -70 km/s relative to the stars and H II regions and presumably arises in a galaxy-scale outflow similar to those seen in most star-forming galaxies at z = 2-3. The other is more unusual in showing a mean redshift of +350 km/s relative to the systemic redshift; possible interpretations include a merging clump, or material ejected by a previous star formation episode and now falling back onto the galaxy, or more simply a chance alignment with a foreground galaxy. In the metal absorption lines, both components only partially cover the OB stars against which they are being viewed. We tentatively associate the redshifted component with the strong damped Lyman alpha line, indicative of a column density N(H I) = (3.0 +/- 0.8) x 10(21) atoms/cm2, and propose that it provides the dust `foreground screen' responsible for the low ratio of far-infrared to UV luminosities of the Cosmic Eye. Compared to other well-studied examples of strongly lensed galaxies, we find that the young stellar population of the Cosmic Eye is essentially indistinguishable from those of the Cosmic Horseshoe and MS 1512-cB58, while the interstellar spectra of all three galaxies are markedly different, attesting to the real complexity of the interplay between starbursts and ambient interstellar matter in young galaxies (abridged).Comment: 14 pages, 6 Figures, Accepted for publication in Monthly Notices of the Royal Astronomical Society after minor revision

New technologies for diagnosing active TB: the VANTDET diagnostic accuracy study

BackgroundTuberculosis (TB) is a devastating disease for which new diagnostic tests are desperately needed.ObjectiveTo validate promising new technologies [namely whole-blood transcriptomics, proteomics, flow cytometry and quantitative reverse transcription-polymerase chain reaction (qRT-PCR)] and existing signatures for the detection of active TB in samples obtained from individuals with suspected active TB.DesignFour substudies, each of which used samples from the biobank collected as part of the interferon gamma release assay (IGRA) in the Diagnostic Evaluation of Active TB study, which was a prospective cohort of patients recruited with suspected TB.SettingSecondary care.ParticipantsAdults aged ≥ 16 years presenting as inpatients or outpatients at 12 NHS hospital trusts in London, Slough, Oxford, Leicester and Birmingham, with suspected active TB.InterventionsNew tests using genome-wide gene expression microarray (transcriptomics), surface-enhanced laser desorption ionisation time-of-flight mass spectrometry/liquid chromatography–mass spectrometry (proteomics), flow cytometry or qRT-PCR.Main outcome measuresArea under the curve (AUC), sensitivity and specificity were calculated to determine diagnostic accuracy. Positive and negative predictive values were calculated in some cases. A decision tree model was developed to calculate the incremental costs and quality-adjusted life-years of changing from current practice to using the novels tests.ResultsThe project, and four substudies that assessed the previously published signatures, measured each of the new technologies and performed a health economic analysis in which the best-performing tests were evaluated for cost-effectiveness. The diagnostic accuracy of the transcriptomic tests ranged from an AUC of 0.81 to 0.84 for detecting all TB in our cohort. The performance for detecting culture-confirmed TB or pulmonary TB was better than for highly probable TB or extrapulmonary tuberculosis (EPTB), but was not high enough to be clinically useful. None of the previously described serum proteomic signatures for active TB provided good diagnostic accuracy, nor did the candidate rule-out tests. Four out of six previously described cellular immune signatures provided a reasonable level of diagnostic accuracy (AUC = 0.78–0.92) for discriminating all TB from those with other disease and latent TB infection in human immunodeficiency virus-negative TB suspects. Two of these assays may be useful in the IGRA-positive population and can provide high positive predictive value. None of the new tests for TB can be considered cost-effective.LimitationsThe diagnostic performance of new tests among the HIV-positive population was either underpowered or not sufficiently achieved in each substudy.ConclusionsOverall, the diagnostic performance of all previously identified ‘signatures’ of TB was lower than previously reported. This probably reflects the nature of the cohort we used, which includes the harder to diagnose groups, such as culture-unconfirmed TB or EPTB, which were under-represented in previous cohorts.Future workWe are yet to achieve our secondary objective of deriving novel signatures of TB using our data sets. This was beyond the scope of this report. We recommend that future studies using these technologies target specific subtypes of TB, specifically those groups for which new diagnostic tests are required.FundingThis project was funded by the Efficacy and Mechanism Evaluation (EME) programme, a MRC and NIHR partnership

Sensors in the stream: the high-frequency wave of the present

New scientific understanding is catalysed by novel technologies that enhance measurement precision, resolution or type, and that provide new tools to test and develop theory. Over the last 50 years, technology has transformed the hydrologic sciences by enabling direct measurements of watershed fluxes (evapotranspiration, streamflow) at time scales and spatial extents aligned with variation in physical drivers. High frequency water quality measurements, increasingly obtained by in-situ water quality sensors, are extending that transformation. Widely available sensors for some physical (temperature) and chemical (conductivity, dissolved oxygen) attributes have become integral to aquatic science, and emerging sensors for nutrients, dissolved CO2, turbidity, algal pigments, and dissolved organic matter are now enabling observations of watersheds and streams at timescales commensurate with their fundamental hydrological, energetic, elemental, and biological drivers. Here we synthesize insights from emerging technologies across a suite of applications, and envision future advances, enabled by sensors, in our ability to understand, predict, and restore watershed and stream systems

CD4+ T cell cytokine responses to the DAR-901 booster vaccine in BCG-primed adults:A randomized, placebo-controlled trial

<div><p>Background</p><p>DAR-901 is an inactivated whole cell tuberculosis booster vaccine, prepared using a new scalable, broth-grown method from the master cell bank of SRL172, a vaccine previously shown to prevent tuberculosis. This study examined whether DAR-901 (a) induces CD4+ T cell cytokine profiles previously proposed as correlates of protection and (b) has a specific vaccine-induced immunological signature compared to BCG or placebo.</p><p>Methods</p><p>We analysed CD4+ T cell cytokine immune responses from 10 DAR-901 recipients, 9 BCG recipients and 9 placebo recipients from the Phase I DAR-901 MDES trial. In that study, HIV-negative, IGRA-negative participants with prior BCG immunization were randomized (double-blind) to receive three intradermal injections of DAR-901 or saline placebo or two injections of saline placebo followed by an intradermal injection of BCG. Antigen-specific functional and phenotypic CD4+ T cell responses along with effector phenotype of responder cells were measured by intracellular cytokine staining.</p><p>Results</p><p>DAR-901 recipients exhibited increased DAR-901 antigen-specific polyfunctional or bifunctional T cell responses compared to baseline. Vaccine specific CD4+ IFNγ, IL2, TNFα and any cytokine responses peaked at 7 days post-dose 3. Th1 responses predominated, with most responder cells exhibiting a polyfunctional effector memory phenotype. BCG induced greater CD4+ T cell responses than placebo while the more modest DAR-901 responses did not differ from placebo. Neither DAR-901 nor BCG induced substantial or sustained Th17 /Th22 cytokine responses.</p><p>Conclusion</p><p>DAR-901, a TB booster vaccine grown from the master cell bank of SRL 172 which was shown to prevent TB, induced low magnitude polyfunctional effector memory CD4+ T cell responses. DAR-901 responses were lower than those induced by BCG, a vaccine that has been shown ineffective as a booster to prevent tuberculosis disease. These results suggest that induction of higher levels of CD4+ cytokine stimulation may not be a critical or pre-requisite characteristic for candidate TB vaccine boosters.</p><p>Trial registration</p><p>ClinicalTrials.gov <a href="https://clinicaltrials.gov/ct2/show/NCT02063555" target="_blank">NCT02063555</a>.</p></div