The lack of well-preserved material upon which to base the paleo-microbiological detection of Plasmodium parasites has prevented extensive documentation of past outbreaks of malaria in Europe. By trapping intact erythrocytes at the time of death, dental pulp has been shown to be a suitable tissue for documenting ancient intraerythrocytic pathogens such as Plasmodium parasites. Total DNA and proteins extracted from 23 dental pulp specimens collected from individuals exhumed from the 9th to 13th century archaeological site in Mariana, Corsica, were analyzed using open-mind paleo-auto-immunohistochemistry and direct metagenomics, Plasmodium-targeting immunochromatography assays. All experiments incorporated appropriate negative controls. Paleo-auto-immunohistochemistry revealed the presence of parasites Plasmodium spp. in the dental pulp of nine teeth. A further immunochromatography assay identified the presence of at least one Plasmodium antigen in nine individuals. The nine teeth, for which the PfHRP-2 antigen specific of P. falciparum was detected, were also positive using paleo-autoimmunohistochemistry and metagenomics. Dental pulp erythrocytes proved to be suitable for the direct paleomicrobiology documentation of malaria in nine individuals buried in medieval Corsica, in agreement with historical data. This provides additional information on the millennial dynamics of Plasmodium spp. in the Mediterranean basin.

Antibody kinetic curves obtained during a viral infection are often fitted using aggregated patient data, hiding the heterogeneity of individual humoral immune responses. Individual antibody responses can be modeled using the Wood equation and grouped according to their profile. Such modeling takes into account several important kinetic parameters, such as the day when antibody detection becomes positive [daypos], the day of the maximal response [daymax], the maximum antibody level [levelmax], and the day when antibody detection becomes negative [dayneg]. Potential associations between these profiles and studied factors can then be tested.

MALDI-TOF is now considered a relevant tool for the identification of arthropods, including lice and fleas. However, the duration and conditions of storage, such as in ethanol, which is frequently used to preserve these ectoparasites, could impede their classification. The purpose of the present study was to assess the stability of MS profiles from Pediculus humanus corporis lice and Ctenocephalides felis fleas preserved in alcohol from one to four years and kinetically submitted to MALDI-TOF MS. A total of 469 cephalothoraxes from lice (n = 170) and fleas (n = 299) were tested. The reproducibility of the MS profiles was estimated based on the log score values (LSVs) obtained for query profiles compared to the reference profiles included in the MS database. Only MS spectra from P. humanus corporis and C. felis stored in alcohol for less than one year were included in the reference MS database. Approximately 75% of MS spectra from lice (75.2%, 94/125) and fleas (74.4%, 122/164) specimens stored in alcohol for 12 to 48 months, queried against the reference MS database, obtained relevant identification. An accurate analysis revealed a significant decrease in the proportion of identification for both species stored for more than 22 months in alcohol. It was hypothesized that incomplete drying was responsible for MS spectra variations. Then, 45 lice and 60 fleas were subjected to longer drying periods from 12 to 24 h. The increase in the drying period improved the proportion of relevant identification for lice (95%) and fleas (80%). This study highlighted that a correct rate of identification by MS could be obtained for lice and fleas preserved in alcohol for up to four years on the condition that the drying period was sufficiently long for accurate identification.

AbstractThe risk of mosquito-borne diseases transmission is moving fast toward temperate climates with the colonization and proliferation of the Asian tiger mosquito vectorAedes albopictusand the rapid and mass transport of passengers returning from tropical regions where the viruses are endemic. The prevention of majorAedes-borne viruses heavily relies on the use of insecticides for vector control, mainly pyrethroids In Europe. High-throughput molecular assays can provide a cost-effective surrogate to phenotypic insecticide resistance assays when mutations have been previously linked to a resistance phenotype. Here, we screened for the spatial distribution ofkdrmutations at a large scale using a two-step approach based on multiplexed amplicon sequencing and an unprecedented collection of field-derived mosquitoes in South of France. We identified the presence of the V1016G allele in 14 sites. The V1016G allele was predominantly found in South-East France close to the Italian border with two additional isolated sites close to Bordeaux and Marmande. All mosquitoes were heterozygous for this mutation and should not be phenotypically resistant to pyrethroid insecticide. Four other mutations were identified in our targeted genomic sequence: I1532T, M1006L, M1586L, M995L. Sequencing a section of maternally inherited mitochondrial genome confirmed that the spread ofAe. albopictusin France originated from founders with haplogroup A1. These findings contribute to the broader understanding of resistance dynamics in Europe and can inform targeted approaches to mitigate the impact of resistance on vector control.

Chemical control of disease vectoring mosquitoes Aedes albopictus and Aedes aegypti is costly, unsustainable, and increasingly ineffective due to the spread of insecticide resistance. The Sterile Insect Technique is a valuable alternative but is limited by slow, error-prone, and wasteful sex-separation methods. Here, we present four Genetic Sexing Strains (two for each Aedes species) based on fluorescence markers linked to the m and M sex loci, allowing for the isolation of transgenic males. Furthermore, we demonstrate how combining these sexing strains enables the production of non-transgenic males. In a mass-rearing facility, 100,000 first instar male larvae could be sorted in under 1.5 h with an estimated 0.01–0.1% female contamination on a single machine. Cost-efficiency analyses revealed that using these strains could result in important savings while setting up and running a mass-rearing facility. Altogether, these Genetic Sexing Strains should enable a major upscaling in control programmes against these important vectors.

Two new bacterial strains, Marseille-P2698T (CSUR P2698 = DSM 103,121) and Marseille-P2260T (CSUR P2260 = DSM 101,844 = SN18), were isolated from human stools by the culturomic method. We used the taxonogenomic approach to fully describe these two new bacterial strains. The Marseille-P2698T strain was a Gram-negative, motile, non-spore-forming, rod-shaped bacterium. The Marseille-P2260T strain was a Gram-positive, motile, spore-forming rod-shaped bacterium. Major fatty acids found in Marseille-P2698T were C15:0 iso (63%), C15:0 anteiso (11%), and C17:0 3-OH iso (8%). Those found in Marseille-P2260T strain were C16:00 (39%), C18:1n9 (16%) and C18:1n7 (14%). Strains Marseille-P2698T and Marseille-P2260T had 16S rRNA gene sequence similarities of 91.50% with Odoribacter laneusT, and of 90.98% and 95.07% with Odoribacter splanchnicusT and Eubacterium sulciT, respectively. The exhibited digital DNA-DNA Hybridization values lower than 20.7%, and Orthologous Average Nucleotide Identity values lower than 73% compared to their closest related bacterial species O. splanchnicusT and E. sulciT respectively. Phenotypic, biochemical, phylogenetic, and genomic results obtained by comparative analyses provided sufficient evidence that both of the two studied strains Marseille-P2698T and Marseille-P2260T are two new bacterial species and new bacterial genera for which the names Culturomica massiliensis gen. nov., sp. nov., and Emergencia timonensis gen. nov., sp. nov. were proposed, respectively.

BackgroundPaleomicrobiological data have clarified that Plasmodium spp. was circulating in the past in southern European populations, which are now devoid of malaria. The aim of this study was to evaluate the efficacy of immunodetection and, more particularly, rapid diagnostic tests (RDT), in order to further assess Plasmodium infections in ancient northern European populations.MethodsA commercially available RDT, PALUTOP® + 4 OPTIMA, which is routinely used to detect malaria, was used to detect Plasmodium antigens from proteins recovered from ancient specimens extracted from 39 dental pulp samples. These samples were collected from 39 individuals who were buried in the sixth century, near the site of the current Palace of Versailles in France. Positive and negative controls were also used. Antigens detected were quantified using chemiluminescence imaging system analysis.ResultsPlasmodium antigens were detected in 14/39 (35.9%) individuals, including Plasmodium vivax antigens in 11 individuals and Plasmodium falciparum antigens co-detected in two individuals, while Pan-Plasmodium antigens were detected in three individuals. Controls all yielded expected results.ConclusionsThe data reported here showed that RDTs are a suitable tool for detecting Plasmodium spp. antigens in ancient dental pulp samples, and demonstrated the existence of malaria in Versailles, France, in the sixth century. Plasmodium vivax, which is regarded as being responsible for an attenuated form of malaria and less deadly forms, was the most prevalent species. This illustrates, for the first time in ancient populations, co-infection with P. falciparum, bringing into question the climate-driven ecosystems prevailing at that time in the Versailles area.

BackgroundThe Republic of Djibouti is a malaria endemic country that was in pre-elimination phase in 2006–2012. From 2013, however, malaria has re-emerged in the country, and its prevalence has been increasing every year. Given the co-circulation of several infectious agents in the country, the assessment of malaria infection based on microscopy or histidine-rich protein 2 (HRP2)-based rapid diagnostic tests (RDT) has shown its limitations. This study, therefore, aimed to assess the prevalence of malaria among febrile patients in Djibouti city using more robust molecular tools.MethodsAll suspected malaria cases reported to be microscopy-positive were randomly sampled (n = 1113) and included in four health structures in Djibouti city over a 4-year period (2018–2021), mainly during the malaria transmission season (January–May). Socio-demographic information was collected, and RDT was performed in most of the included patients. The diagnosis was confirmed by species-specific nested polymerase chain reaction (PCR). Data were analysed using Fisher’s exact test and kappa statistics.ResultsIn total, 1113 patients with suspected malaria and available blood samples were included. PCR confirmed that 788/1113 (70.8%) were positive for malaria. Among PCR-positive samples, 656 (83.2%) were due to Plasmodium falciparum, 88 (11.2%) Plasmodium vivax, and 44 (5.6%) P. falciparum/P. vivax mixed infections. In 2020, P. falciparum infections were confirmed by PCR in 50% (144/288) of negative RDTs. After the change of RDT in 2021, this percentage decreased to 17%. False negative RDT results were found more frequently (P < 0.05) in four districts of Djibouti city (Balbala, Quartier 7, Quartier 6, and Arhiba). Malaria occurred less frequently in regular bed net users than in non-users (odds ratio [OR]: 0.62, 95% confidence interval [CI]: 0.42–0.92).ConclusionsThe present study confirmed the high prevalence of falciparum malaria and, to a lesser extent, vivax malaria. Nevertheless, 29% of suspected malaria cases were misdiagnosed by microscopy and/or RDT. There is a need to strengthen the capacity for diagnosis by microscopy and to evaluate the possible role of P. falciparum hrp2 gene deletion, which leads to false negative cases of P. falciparum.

International travellers frequently acquire infectious diseases whilst travelling, yet relatively little is known about the impact and economic burden of these illnesses on travellers. We conducted a prospective exploratory costing study on adult returning travellers with falciparum malaria, dengue, chikungunya or Zika virus. Patients were recruited in eight Travel and Tropical Medicine clinics between June 2016 and March 2020 upon travellers' first contact with the health system in their country of residence. The patients were presented with a structured 52-question self-administered questionnaire after full recovery to collect information on patients' healthcare utilization and out-of-pocket costs both in the destination and home country, and about income and other financial losses due to the illness. A total of 134 patients participated in the study (malaria, 66; dengue, 51; chikungunya, 8; Zika virus, 9; all fully recovered; median age 40; range 18-72years). Prior to travelling, 42% of patients reported procuring medical evacuation insurance. Across the four illnesses, only 7% of patients were hospitalized abroad compared with 61% at home. Similarly, 15% sought ambulatory services whilst abroad compared with 61% at home. The average direct out-of-pocket hospitalization cost in the destination country (USD $2236; range: $108-$5160) was higher than the direct out-of-pocket ambulatory cost in the destination country (USD $327; range: $0-$1560), the direct out-of-pocket hospitalization cost at home (USD $35; range: $0-$120) and the direct out-of-pocket ambulatory costs at home (US$45; range: $0-$192). Respondents with dengue or malaria lost a median of USD $570 (Interquartile range [IQR] 240-1140) and USD $240 (IQR 0-600), respectively, due to their illness, whilst those with chikungunya and Zika virus lost a median of USD $2400 (IQR 1200-3600) and USD $1500 (IQR 510-2625), respectively. Travellers often incur significant costs due to travel-acquired diseases. Further research into the economic impact of these diseases on travellers should be conducted.