Disruption of a Plasmodium falciparum patatin-like phospholipase delays male gametocyte exflagellation.

An essential process in transmission of the malaria parasite to the Anopheles vector is the conversion of mature gametocytes into gametes within the mosquito gut, where they egress from the red blood cell (RBC). During egress, male gametocytes undergo exflagellation, leading to the formation of eight haploid motile microgametes, while female gametes retain their spherical shape. Gametocyte egress depends on sequential disruption of the parasitophorous vacuole membrane and the host cell membrane. In other life cycle stages of the malaria parasite, phospholipases have been implicated in membrane disruption processes during egress, however their importance for gametocyte egress is relatively unknown. Here, we performed comprehensive functional analyses of six putative phospholipases for their role during development and egress of Plasmodium falciparum gametocytes. We localize two of them, the prodrug activation and resistance esterase (PF3D7_0709700) and the lysophospholipase 1 (PF3D7_1476700), to the parasite plasma membrane. Subsequently, we show that disruption of most of the studied phospholipase genes does neither affect gametocyte development nor egress. The exception is the putative patatin-like phospholipase 3 (PF3D7_0924000), whose gene deletion leads to a delay in male gametocyte exflagellation, indicating an important, albeit not essential, role of this enzyme in male gametogenesis.

Autoimmunity Against Surfactant Protein B Is Associated with Pneumonitis During Checkpoint Blockade.

Rationale: Immune checkpoint inhibitor (ICI)-related pneumonitis is a serious autoimmune event affecting as many as 20% of patients with non-small-cell lung cancer (NSCLC), yet the factors underpinning its development in some patients and not others are poorly understood. Objectives: To investigate the role of autoantibodies and autoreactive T cells against surfactant-related proteins in the development of pneumonitis. Methods: The study cohort consisted of patients with NSCLC who provided blood samples before and during ICI treatment. Serum was used for proteomics analyses and to detect autoantibodies present during pneumonitis. T-cell stimulation assays and single-cell RNA sequencing were performed to investigate the specificity and functionality of peripheral autoreactive T cells. The findings were confirmed in a validation cohort comprising patients with NSCLC and patients with melanoma. Measurements and Main Results: Across both cohorts, patients in whom pneumonitis developed had higher pretreatment levels of immunoglobulin G autoantibodies targeting surfactant protein (SP)-B. At the onset of pneumonitis, these patients also exhibited higher frequencies of CD4+ IFN-γ-positive SP-B-specific T cells and expanding T-cell clonotypes recognizing this protein, accompanied by a proinflammatory serum proteomic profile. Conclusions: Our data suggest that the cooccurrence of SP-B-specific immunoglobulin G autoantibodies and CD4+ T cells is associated with the development of pneumonitis during ICI therapy. Pretreatment levels of these antibodies may represent a potential biomarker for an increased risk of developing pneumonitis, and on-treatment levels may provide a diagnostic aid.

A national surveillance system for continuous monitoring of blood transfusion safety: German haemovigilance data.

BACKGROUND AND OBJECTIVES: Haemovigilance (HV) systems aim to improve transfusion outcomes in patients and donor safety. An important question for blood regulators is how to ensure an effective HV system. MATERIALS AND METHODS: We retrospectively analysed the HV reports submitted to Paul-Ehrlich-Institut over the last two decades. RESULTS: Between 2011 and 2020, 50.86 million units of blood components were used, and 8931 suspected serious donor and recipient adverse reactions (SARs), 874 serious adverse events (SAEs) and 12,073 donor look-backs were reported. Following implementation of specific risk-minimization measures (RMMs) between 2000 and 2010, SAR reporting rates decreased for transfusion-transmitted viral infections (TTVIs), transfusion-related acute lung injury (TRALI) and transfusion-transmitted bacterial infections (TTBIs), while increasing for other serious adverse transfusion reactions. Within this decade, the overall blood component use decreased. CONCLUSION: Long-term data collection forms the basis to establish trends and changes in reporting and to evaluate the effect of RMM. Standardized criteria for reaction types, seriousness and imputability assessments and availability of a denominator are important elements. Central data collection and independent assessment allow for monitoring HV data in a nationwide context over time. Stakeholder involvement and transparent feedback on the benefit of RMM will help to achieve the objectives of HV.

Model-guided development of an evolutionarily stable yeast chassis.

First-principle metabolic modelling holds potential for designing microbial chassis that are resilient against phenotype reversal due to adaptive mutations. Yet, the theory of model-based chassis design has rarely been put to rigorous experimental test. Here, we report the development of Saccharomyces cerevisiae chassis strains for dicarboxylic acid production using genome-scale metabolic modelling. The chassis strains, albeit geared for higher flux towards succinate, fumarate and malate, do not appreciably secrete these metabolites. As predicted by the model, introducing product-specific TCA cycle disruptions resulted in the secretion of the corresponding acid. Adaptive laboratory evolution further improved production of succinate and fumarate, demonstrating the evolutionary robustness of the engineered cells. In the case of malate, multi-omics analysis revealed a flux bypass at peroxisomal malate dehydrogenase that was missing in the yeast metabolic model. In all three cases, flux balance analysis integrating transcriptomics, proteomics and metabolomics data confirmed the flux re-routing predicted by the model. Taken together, our modelling and experimental results have implications for the computer-aided design of microbial cell factories.

Identification of the 3-amino-3-carboxypropyl (acp) transferase enzyme responsible for acp3U formation at position 47 in Escherichia coli tRNAs.

tRNAs from all domains of life contain modified nucleotides. However, even for the experimentally most thoroughly characterized model organism Escherichia coli not all tRNA modification enzymes are known. In particular, no enzyme has been found yet for introducing the acp3U modification at position 47 in the variable loop of eight E. coli tRNAs. Here we identify the so far functionally uncharacterized YfiP protein as the SAM-dependent 3-amino-3-carboxypropyl transferase catalyzing this modification and thereby extend the list of known tRNA modification enzymes in E. coli. Similar to the Tsr3 enzymes that introduce acp modifications at U or m1Ψ nucleotides in rRNAs this protein contains a DTW domain suggesting that acp transfer reactions to RNA nucleotides are a general function of DTW domain containing proteins. The introduction of the acp3U-47 modification in E. coli tRNAs is promoted by the presence of the m7G-46 modification as well as by growth in rich medium. However, a deletion of the enzymes responsible for the modifications at position 46 and 47 in the variable loop of E. coli tRNAs did not lead to a clearly discernible phenotype suggesting that these two modifications play only a minor role in ensuring the proper function of tRNAs in E. coli.

Experimental Parasite Infection Causes Genome-Wide Changes in DNA Methylation.

Parasites are arguably among the strongest drivers of natural selection, constraining hosts to evolve resistance and tolerance mechanisms. Although, the genetic basis of adaptation to parasite infection has been widely studied, little is known about how epigenetic changes contribute to parasite resistance and eventually, adaptation. Here, we investigated the role of host DNA methylation modifications to respond to parasite infections. In a controlled infection experiment, we used the three-spined stickleback fish, a model species for host-parasite studies, and their nematode parasite Camallanus lacustris. We showed that the levels of DNA methylation are higher in infected fish. Results furthermore suggest correlations between DNA methylation and shifts in key fitness and immune traits between infected and control fish, including respiratory burst and functional trans-generational traits such as the concentration of motile sperm. We revealed that genes associated with metabolic, developmental, and regulatory processes (cell death and apoptosis) were differentially methylated between infected and control fish. Interestingly, genes such as the neuropeptide FF receptor 2 and the integrin alpha 1 as well as molecular pathways including the Th1 and Th2 cell differentiation were hypermethylated in infected fish, suggesting parasite-mediated repression mechanisms of immune responses. Altogether, we demonstrate that parasite infection contributes to genome-wide DNA methylation modifications. Our study brings novel insights into the evolution of vertebrate immunity and suggests that epigenetic mechanisms are complementary to genetic responses against parasite-mediated selection.

The time is ripe for functional genomics: Can epigenetic changes mediate reproductive timing?

Populations are under strong selection to match reproductive timing with favourable environmental conditions. This becomes particularly important and challenging with increasing interannual environmental variability. Adjusting reproductive timing requires the ability to sense and interpret relevant environmental cues, while responding flexibly to their interannual variation. For instance, in seasonal species, reproductive timing is often dependent on photoperiod and temperature. Although many genes influencing the timing of reproduction have been identified, far less attention has been paid to the gene-regulatory cascades orchestrating these complex gene-environment interactions. In a From the Cover article in this issue of Molecular Ecology, Lindner, Laine, et al. (2021) addressed this knowledge gap by investigating the role of DNA methylation in mediating reproductive timing in the seasonally breeding great tit (Parus major). Using a clever blood sampling design, they investigated genome-wide DNA methylation changes following individual female birds across multiple reproductive stages. This approach revealed 10 candidate genes with a strong correlation between promoter methylation and reproductive status. Some of these genes are known to be involved in reproductive timing (e.g., MYLK-like or NR5A1), yet for others this function was previously unknown (Figure 1). Interestingly, NR5A1 is a key transcription factor, which may affect other genes that are part of the same regulatory network. The findings of Lindner, Laine, et al. (2021) provide a strong case for studying DNA methylation to uncover how gene-environment interactions influence important life-history traits, such as reproductive timing.

Individual variability and versatility in an eco-evolutionary model of avian migration.

Seasonal migration is a complex and variable behaviour with the potential to promote reproductive isolation. In Eurasian blackcaps (Sylvia atricapilla), a migratory divide in central Europe separating populations with southwest (SW) and southeast (SE) autumn routes may facilitate isolation, and individuals using new wintering areas in Britain show divergence from Mediterranean winterers. We tracked 100 blackcaps in the wild to characterize these strategies. Blackcaps to the west and east of the divide used predominantly SW and SE directions, respectively, but close to the contact zone many individuals took intermediate (S) routes. At 14.0° E, we documented a sharp transition from SW to SE migratory directions across only 27 (10-86) km, implying a strong selection gradient across the divide. Blackcaps wintering in Britain took northwesterly migration routes from continental European breeding grounds. They originated from a surprisingly extensive area, spanning 2000 km of the breeding range. British winterers bred in sympatry with SW-bound migrants but arrived 9.8 days earlier on the breeding grounds, suggesting some potential for assortative mating by timing. Overall, our data reveal complex variation in songbird migration and suggest that selection can maintain variation in migration direction across short distances while enabling the spread of a novel strategy across a wide range.

Psychometric characterization of incidental feedback sources during grasping with a hand prosthesis.

BACKGROUND: A prosthetic system should ideally reinstate the bidirectional communication between the user's brain and its end effector by restoring both motor and sensory functions lost after an amputation. However, current commercial prostheses generally do not incorporate somatosensory feedback. Even without explicit feedback, grasping using a prosthesis partly relies on sensory information. Indeed, the prosthesis operation is characterized by visual and sound cues that could be exploited by the user to estimate the prosthesis state. However, the quality of this incidental feedback has not been objectively evaluated. METHODS: In this study, the psychometric properties of the auditory and visual feedback of prosthesis motion were assessed and compared to that of a vibro-tactile interface. Twelve able-bodied subjects passively observed prosthesis closing and grasping an object, and they were asked to discriminate (experiment I) or estimate (experiment II) the closing velocity of the prosthesis using visual (VIS), acoustic (SND), or combined (VIS + SND) feedback. In experiment II, the subjects performed the task also with a vibrotactile stimulus (VIB) delivered using a single tactor. The outcome measures for the discrimination and estimation experiments were just noticeable difference (JND) and median absolute estimation error (MAE), respectively. RESULTS: The results demonstrated that the incidental sources provided a remarkably good discrimination and estimation of the closing velocity, significantly outperforming the vibrotactile feedback. Using incidental sources, the subjects could discriminate almost the minimum possible increment/decrement in velocity that could be commanded to the prosthesis (median JND < 2% for SND and VIS + SND). Similarly, the median MAE in estimating the prosthesis velocity randomly commanded from the full working range was also low, i.e., approximately 5% in SND and VIS + SND. CONCLUSIONS: Since the closing velocity is proportional to grasping force in state-of-the-art myoelectric prostheses, the results of the present study imply that the incidental feedback, when available, could be usefully exploited for grasping force control. Therefore, the impact of incidental feedback needs to be considered when designing a feedback interface in prosthetics, especially since the quality of estimation using supplemental sources (e.g., vibration) can be worse compared to that of the intrinsic cues.

Detection and Correlation of Single and Concomitant TP53, PTEN, and CDKN2A Alterations in Gliomas.

Gliomas are the most frequent primary tumors of central nervous system and represent a heterogeneous group of tumors that originates from the glial cells. TP53, PTEN, and CDKN2A are important tumor suppressor genes that encode proteins involved in sustaining cellular homeostasis by different signaling pathways. Though genetic alterations in these genes play a significant role in tumorigenesis, few studies are available regarding the incidence and relation of concomitant TP53, PTEN, and CDKN2A alterations in gliomas. The purpose of this study was to evaluate the occurrence of mutation and deletion in these genes, through single-strand conformational polymorphism, array-comparative genomic hybridization, and fluorescence in situ hybridization techniques, in 69 gliomas samples. Molecular results demonstrated a significant higher prevalence of TP53, PTEN, and CDKN2A alterations in astrocytoma than other tumor subtypes, and heterozygous deletion was the most frequent event. In addition, a significant association was observed between TP53 and CDKN2A alterations (p = 0.0424), which tend to coexist in low grade astrocytomas (5/46 cases (10.9%)), suggesting that they are early events in development of these tumors, and PTEN and CDKN2A deletions (p = 0.0022), which occurred concomitantly in 9/50 (18%) patients, with CDKN2A changes preceding PTEN deletions, present preferably in high-grade gliomas.