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Abstract Zebrafish is widely adopted as a grafting model for studying human development and diseases. Current zebrafish xenotransplantations are performed using embryo recipients, as the adaptive immune system, responsible for host versus graft rejection, only reaches maturity at juvenile stage. However, transplanted primary human hematopoietic stem/progenitor cells (HSC) rapidly disappear even in zebrafish embryos, suggesting that another barrier to transplantation exists before the onset of adaptive immunity. Here, using a labelled macrophage zebrafish line, we demonstrated that engraftment of human HSC induces a massive recruitment of macrophages which rapidly phagocyte transplanted cells. Macrophages depletion, by chemical or pharmacological treatments, significantly improved the uptake and survival of transplanted cells, demonstrating the crucial implication of these innate immune cells for the successful engraftment of human cells in zebrafish. Beyond identifying the reasons for human hematopoietic cell engraftment failure, this work images the fate of human cells in real time over several days in macrophage-depleted zebrafish embryos.

Recently, many studies demonstrated the fundamental role of the tumour microenvironment (TME) in cancer progression. Here, we describe a method to visualize in 3D the behaviour of tumours in zebrafish embryos. We highlight two major actors of the TME, macrophages and vessels. This valuable tool is transposable to Patient-Derived Xenograft imaging in order to predict the fate of malignant tumours according to the dynamics of their TME.

Abstract The inability to control cell proliferation results in the formation of tumors in many multicellular lineages. Nonetheless, little is known about the extent of conservation of the biological traits and ecological factors that promote or inhibit tumorigenesis across the metazoan tree. Particularly, changes in food availability have been linked to increased cancer incidence in humans, as an outcome of evolutionary mismatch. Here, we apply evolutionary oncology principles to test whether food availability, regardless of the multicellular lineage considered, has an impact on tumorigenesis. We used two phylogenetically unrelated model systems, the cnidarian Hydra oligactis and the fish Danio rerio, to investigate the impact of resource availability on tumor occurrence and progression. Individuals from healthy and tumor-prone lines were placed on four diets that differed in feeding frequency and quantity. For both models, frequent overfeeding favored tumor emergence, while lean diets appeared more protective. In terms of tumor progression, high food availability promoted it, whereas low resources controlled it, but without having a curative effect. We discuss our results in light of current ideas about the possible conservation of basic processes governing cancer in metazoans (including ancestral life history trade-offs at the cell level) and in the framework of evolutionary medicine.

Abstract Background Dengue fever is a major public health concern in Cambodia, with thousands of cases every year in urban, suburban and rural areas of the country. The main vector of dengue fever in Cambodia is Aedes aegypti. The organophosphate larvicide temephos and adulticides belonging to the pyrethroid family have been widely used for decades by public health authorities to fight dengue vectors, but resistance of Ae. aegypti to these insecticides has been previously described for Cambodia. Methods In order to adapt the vector control strategy presently used in Cambodia, we tested 14 adulticides belonging to the carbamate, organochlorine, organophosphate, and pyrethroid insecticide families and three larvicides [temephos, spinosad and Bacillus thuringiensis ser. israelensis (Bti)] belonging to three different insecticide families (organophosphates, spinosyns and entomopathogenic bacteria). The standard procedures used here to test the adults and larvae of an Ae. aegypti population from Phnom Penh followed World Health Organization guidelines. Results For adults, high mortality rates were observed with carbamate, organophosphate and organochlorine (with the exception of dichlorodiphenyltrichloroethane) insecticides (i.e. between 87.6 and 100%), while low mortality rates were observed with all of the tested pyrethroid insecticides (i.e. between 1 and 35%). For larvae, no resistance against Bti was detected [resistance ratio (RR90

Summary: Background: Encephalitis is a worldwide public health issue, with a substantially high burden among children in southeast Asia. We aimed to determine the causes of encephalitis in children admitted to hospitals across the Greater Mekong region by implementing a comprehensive state-of-the-art diagnostic procedure harmonised across all centres, and identifying clinical characteristics related to patients’ conditions. Methods: In this multicentre, observational, prospective study of childhood encephalitis, four referral hospitals in Cambodia, Vietnam, Laos, and Myanmar recruited children (aged 28 days to 16 years) who presented with altered mental status lasting more than 24 h and two of the following minor criteria: fever (within the 72 h before or after presentation), one or more generalised or partial seizures (excluding febrile seizures), a new-onset focal neurological deficit, cerebrospinal fluid (CSF) white blood cell count of 5 per mL or higher, or brain imaging (CT or MRI) suggestive of lesions of encephalitis. Comprehensive diagnostic procedures were harmonised across all centres, with first-line testing was done on samples taken at inclusion and results delivered within 24 h of inclusion for main treatable causes of disease and second-line testing was done thereafter for mostly non-treatable causes. An independent expert medical panel reviewed the charts and attribution of causes of all the included children. Using multivariate analyses, we assessed risk factors associated with unfavourable outcomes (ie, severe neurological sequelae and death) at discharge using data from baseline and day 2 after inclusion. This study is registered with ClinicalTrials.gov, NCT04089436, and is now complete. Findings: Between July 28, 2014, and Dec 31, 2017, 664 children with encephalitis were enrolled. Median age was 4·3 years (1·8–8·8), 295 (44%) children were female, and 369 (56%) were male. A confirmed or probable cause of encephalitis was identified in 425 (64%) patients: 216 (33%) of 664 cases were due to Japanese encephalitis virus, 27 (4%) were due to dengue virus, 26 (4%) were due to influenza virus, 24 (4%) were due to herpes simplex virus 1, 18 (3%) were due to Mycobacterium tuberculosis, 17 (3%) were due to Streptococcus pneumoniae, 17 (3%) were due to enterovirus A71, 74 (9%) were due to other pathogens, and six (1%) were due to autoimmune encephalitis. Diagnosis was made within 24 h of admission to hospital for 83 (13%) of 664 children. 119 (18%) children had treatable conditions and 276 (42%) had conditions that could have been preventable by vaccination. At time of discharge, 153 (23%) of 664 children had severe neurological sequelae and 83 (13%) had died. In multivariate analyses, risk factors for unfavourable outcome were diagnosis of M tuberculosis infection upon admission (odds ratio 3·23 [95% CI 1·04–10·03]), coma on day 2 (2·90 [1·78–4·72]), supplementary oxygen requirement (1·89 [1·25–2·86]), and more than 1 week duration between symptom onset and admission to hospital (3·03 [1·68–5·48]). At 1 year after inclusion, of 432 children who were discharged alive from hospital with follow-up data, 24 (5%) had died, 129 (30%) had neurological sequelae, and 279 (65%) had completely recovered. Interpretation: In southeast Asia, most causes of childhood encephalitis are either preventable or treatable, with Japanese encephalitis virus being the most common cause. We provide crucial information that could guide public health policy to improve diagnostic, vaccination, and early therapeutic guidelines on childhood encephalitis in the Greater Mekong region. Funding: Institut Pasteur, Institut Pasteur International Network, Fondation Merieux, Aviesan Sud, INSERM, Wellcome Trust, Institut de Recherche pour le Développement (IRD), and Fondation Total.

Rabies is endemic in Cambodia. For exposed humans, post-exposure prophylaxis (PEP) is very effective in preventing this otherwise fatal disease. The Institut Pasteur du Cambodge (IPC) in Phnom Penh was the primary distributor of PEP in Cambodia until 2018. Since then, and to increase distribution of PEP, two new centers have been opened by IPC in the provinces of Battambang and Kampong Cham. Data on bitten patients, who sometimes bring the head of the biting animal for rabies analyses, have been recorded by IPC since 2000. However, human cases are not routinely recorded in Cambodia, making it difficult to establish a human burden of disease and generate a risk map of dog bites to inform the selection of future PEP center locations in high-risk areas. Our aim was to assess the impact of accessibility to rabies centers on the yearly rate of PEP patients in the population and generate a risk map to identify the locations where new centers would be the most beneficial to the Cambodian population. To accomplish this, we used spatio-temporal Bayesian regression models with the number of PEP patients as the outcome. The primary exposure variable considered was travel time to the nearest IPC center. Secondary exposure variables consisted of travel time to a provincial capital and urban proportion of the population. Between 2000 and 2016, a total of 293,955 PEP patient records were identified. Our results showed a significant negative association between travel time to IPC and the rate of PEP patients: an increase in one hour travel time from the living location to IPC PEP centers leads to a reduction in PEP rate of 70% to 80%. Five provinces were identified as the most efficient locations for future centers to maximize PEP accessibility: Banteay Meanchey, Siem Reap, Takeo, Kampot and Svay Rieng. Adding a PEP center in every provincial capital would increase the proportion of Cambodians living within 60 minutes of a PEP center from 26.6% to 64.9%, and living within 120 minutes from 52.8% to 93.3%, which could save hundreds of lives annually.

The formation of biofilms in dairy processing plants can reduce equipment efficiency, contribute to surface deterioration, and contaminate dairy products by releasing the microorganisms they contain, which may cause spoilage or disease. However, a more representative identification of microbial communities and physico-chemical characterization requires to detach and recover adequately the entire biofilm from the surface. The aim of this study is to develop an efficient technique for in-plant biofilm sampling by growing a strain of Pseudomonas azotoformans PFl1A on stainless-steel surface in a dynamic CDC biofilm reactor system using tryptic soy broth (TSB) and milk as growth media. Different techniques, namely, swabbing, scraping, sonic brushing, synthetic sponge, and sonicating synthetic sponge were used and the results were compared to a standard ASTM International method using ultrasonication. Their efficiencies were evaluated by cells enumeration and scanning electron microscopy. The maximum total viable counts of 8.65 ± 0.06, 8.75 ± 0.08, and 8.71 ± 0.09 log CFU/cm2 were obtained in TSB medium using scraping, synthetic sponge, and sonicating synthetic sponge, respectively, which showed no statistically significant differences with the standard method, ultrasonication (8.74 ± 0.02 log CFU/cm2). However, a significantly (p < 0.05) lower cell recovery of 8.57 ± 0.10 and 8.60 ± 0.00 log CFU/cm2 compared to ultrasonication were achieved for swabbing and sonic brushing, respectively. Furthermore, scanning electron microscopy showed an effective removal of biofilms by sonic brushing, synthetic sponge, and sonicating synthetic sponge; However, only the latter two methods guaranteed a superior release of bacterial biofilm into suspension. Nevertheless, a combination of sonication and synthetic sponge ensured dislodging of sessile cells from surface crevices. The results suggest that a sonicating synthetic sponge could be a promising method for biofilm recovery in processing plants, which can be practically used in the dairy industries as an alternative to ultrasonication.

Southern France, like the rest of the world, is facing the emergence of diseases affecting plants, animals and humans, of which causative agents (viruses, parasites, bacteria) are transmitted by arthropod vectors. Global changes are accelerating the emergence and spread of these diseases. After presenting some examples related to vectors of yellow fever and dengue viruses (Aedes aegypti and Ae. albopictus), Crimean-Congo hemorrhagic fever (Hyalomma marginatum), Bluetongue (Culicoides sp.), and the phytopathogen Xylella fastidiosa (Hemiptera spp.), we will discuss what are the intrinsic and extrinsic factors that make an arthropod a vector in a given place and at a given time. We also propose some thoughts regarding these emergences, possible scenarios for their evolution and some recommendations for the future.

Germline mutations that activate genes in the canonical RAS/MAPK signaling pathway are responsible for rare human developmental disorders known as RASopathies. Here, we analyzed the molecular determinants of Costello syndrome (CS) using a mouse model expressing HRAS p.G12S, patient skin fibroblasts, hiPSC-derived human cardiomyocytes, a HRAS p.G12V zebrafish model, and human fibroblasts expressing lentiviral constructs carrying HRAS p.G12S or HRAS p.G12A mutations. The findings revealed alteration of mitochondrial proteostasis and defective oxidative phosphorylation in the heart and skeletal muscle of CS mice that were also found in the cell models of the disease. The underpinning mechanisms involved the inhibition of the AMPK signaling pathway by mutant forms of HRAS, leading to alteration of mitochondrial proteostasis and bioenergetics. Pharmacological activation of mitochondrial bioenergetics and quality control restored organelle function in HRAS p.G12A and p.G12S cell models, reduced left ventricle hypertrophy in CS mice, and diminished the occurrence of developmental defects in the CS zebrafish model. Collectively, these findings highlight the importance of mitochondrial proteostasis and bioenergetics in the pathophysiology of RASopathies and suggest that patients with CS may benefit from treatment with mitochondrial modulators.

Recent studies have highlighted the large diversity of malaria parasites infecting African great apes (subgenus Laverania) and their strong host specificity. Although the existence of genetic incompatibilities preventing the cross-species transfer may explain host specificity, the existence of vectors with a high preference for a determined host represents another possibility. To test this hypothesis, we undertook a 15-mo-long longitudinal entomological survey in two forest regions of Gabon, where wild apes live, at different heights under the canopy. More than 2,400 anopheline mosquitoes belonging to 18 species were collected. Among them, only three species of Anopheles were found infected with ape Plasmodium: Anopheles vinckei, Anopheles moucheti, and Anopheles marshallii Their role in transmission was confirmed by the detection of the parasites in their salivary glands. Among these species, An. vinckei showed significantly the highest prevalence of infection and was shown to be able to transmit parasites of both chimpanzees and gorillas. Transmission was also shown to be conditioned by seasonal factors and by the heights of capture under the canopy. Moreover, human landing catches of sylvan Anopheles demonstrated the propensity of these three vector species to feed on humans when available. Our results suggest therefore that the strong host specificity observed in the Laveranias is not linked to a specific association between the vertebrate host and the vector species and highlight the potential role of these vectors as bridge between apes and humans.

Whether the malaria parasite Plasmodium falciparum can manipulate mosquito host choice in ways that enhance parasite transmission toward humans is unknown. We assessed the influence of P. falciparum on the blood-feeding behaviour of three of its major vectors (Anopheles coluzzii, An. gambiae and An. arabiensis) in Burkina Faso. Host preference assays using odour-baited traps revealed no effect of infection on mosquito long-range anthropophily. However, the identification of the blood meal origin of mosquitoes showed that females carrying sporozoites, the mature transmissible stage of the parasite, displayed a 24% increase in anthropophagy compared to both females harbouring oocysts, the parasite immature stage, and uninfected individuals. Using a mathematical model, we further showed that this increased anthropophagy in infectious females resulted in a 250% increase in parasite transmission potential, everything else being equal. This important epidemiological consequence highlights the importance of vector control tools targeting infectious females.

Pathogens, which have recently colonized a new host species or new populations of the same host, are interesting models for understanding how populations may evolve in response to novel environments. During its colonization of South America from Africa, Plasmodium falciparum, the main agent of malaria, has been exposed to new conditions in distinctive new human populations (Amerindian and populations of mixed origins) that likely exerted new selective pressures on the parasite's genome. Among the genes that might have experienced strong selective pressures in response to these environmental changes, the eba genes (erythrocyte-binding antigens genes), which are involved in the invasion of the human red blood cells, constitute good candidates. In this study, we analysed, in South America, the polymorphism of three eba genes (eba-140, eba-175, eba-181) and compared it to the polymorphism observed in African populations. The aim was to determine whether these genes faced selective pressures in South America distinct from what they experienced in Africa. Patterns of genetic variability of these genes were compared to the patterns observed at two housekeeping genes (adsl and serca) and 272 SNPs to separate adaptive effects from demographic effects. We show that, conversely to Africa, eba-140 seemed to be under stronger diversifying selection in South America than eba-175. In contrast, eba-181 did not show any sign of departure from neutrality. These changes in the patterns of selection on the eba genes could be the consequence of changes in the host immune response, the host receptor polymorphisms and/or the ability of the parasite to silence or express differentially its invasion proteins.

In Cambodia, dengue outbreaks occur each rainy season (May–October) but vary in magnitude. Using national surveillance data, we designed a tool that can predict 90% of the variance in peak magnitude by April, when typically