Parasites & Vectors最新文献

Background: One of the alternatives for controlling malaria is using mosquito symbiotic bacteria to reduce Plasmodium transmission. Species of Serratia, a genus of the Enterobacteriaceae family, have been isolated from the midgut of mosquitoes and are commonly found in water, soil and plant surfaces. These bacteria have shown great promise in blocking the transmission of Plasmodium in mosquitoes. The aim of this study was to isolate and characterize the genus Serratia within the Anopheles gambiae complex from Burkina Faso.

Methods: Mosquitoes were collected in three field sites located in Houet Province in western Burkina Faso (Dioulassoba, Vallée du Kou and Soumousso), transported to the laboratory and identified morphologically. The salivary gland, midgut, spermatheca, ovary of females and testis of males were dissected and their contents ground up. Different species of Serratia were identified by PCR targeting of the luxS gene of Serratia, followed by 16S ribosomal RNA (rRNA) sequencing.

Results: Molecular analyses identified the isolates as belonging to the genus Serratia, and phylogenetic reconstruction revealed that these strains are highly similar to one another but distinct from Serratia strains previously reported in neighboring countries such as Ghana and Nigeria. The overall prevalence of Serratia among malaria vectors was 13.3%. This prevalence varied according to the development stage of mosquitoes, locality of origin and mosquito organ. Only one Anopheles coluzzii mosquito was co-infected with Serratia and Plasmodium falciparum.

Conclusions: The results of this study support the presence of Serratia spp. in wild mosquitoes from Burkina Faso, we well as their potential use in malaria control.

Background: The introduction of next-generation insecticide-treated nets (ITNs) in Burkina Faso aims to mitigate pyrethroid resistance in malaria vectors. This study evaluated the impact of different ITN types on phenotypic resistance and kdr mutation frequencies in Anopheles gambiae sensus lacto (s.l.) populations across three health districts over 3 years.

Methods: Annual mosquito collections were conducted in Banfora (where pyrethroid-chlorfenapyr nets had been distributed), Gaoua (pyrethroid-only ITNs) and Orodara (pyrethroid-piperonyl butoxide [PBO] ITNs). Two populations were analysed: adult females collected directly from the field and those reared from field-collected larvae. World Health Organization (WHO) susceptibility bioassays measured 24-h mortality after exposure to 1×, 5× and 10× concentrations of deltamethrin and alphacypermethrin, with and without pre-exposure to piperonyl butoxide. Frequencies of kdr mutations L995F and L995S were determined by polymerase chain reaction (PCR).

Results: High-intensity resistance was observed in each study district, with mortality consistently below 45% and not reaching WHO thresholds even at 10× doses. PBO increased mortality, indicating metabolic resistance, but failed to restore full susceptibility. L995F predominated across all districts, years and mosquito populations. L995S remained low and variable. Pyr-only nets were associated with rising L995F frequencies and lower mortality in resistance assays. Pyrethroid (Pyr)-chlorfenapyr (CFR) nets improved mortality in resistance assays without increasing kdr prevalence. Pyr-PBO nets showed partial and inconsistent efficacy, with mosquitoes having mixed patterns in resistance assays. Similar patterns between field and laboratory-reared populations were observed.

Conclusions: ITN type strongly influenced resistance dynamics. Dual-active ingredient (AI) nets, particularly Pyr-CFR, appear more effective in managing resistance. Integrated resistance management combining ITN rotation, routine monitoring and complementary interventions is essential to preserve vector control efficacy.

Background: Helminth infections continue to pose major challenges in human and veterinary medicine, with additional complications arising from the emergence of anthelmintic resistance. Anisakis simplex sensu stricto (A. simplex s.s.), a zoonotic nematode transmitted through the consumption of fish, is of growing concern due to its allergenic potential and clinical relevance. However, the molecular mechanisms underlying the parasite's response to anthelmintic treatment remain poorly defined.

Methods: Third-stage larvae (L3) of A. simplex s.s. were exposed to three widely used anthelmintics: albendazole (ALB), ivermectin (IVC) and pyrantel (PYR). High-throughput RNA sequencing was combined with differential gene expression, multivariate alternative splicing analysis (Replicate Multivariate Analysis of Transcript Splicing [rMATS] v3.2.5 computational tool) and single nucleotide variant (SNV) profiling with Oxford Nanopore sequencing. Drug-specific effects were assessed across protein-coding genes, long non-coding RNAs (lncRNAs) and splicing events.

Results: Distinct transcriptomic features, including splicing and sequence variants, were observed across treatments, with ALB primarily altering the expression of cuticle-associated genes, IVC inducing extensive alternative splicing in immune-related pathways and PYR exposure linked to widespread SNVs in neuronal projection and metabolic genes. Significant splicing events included exon skipping in the trehalase gene (ALB) and combined skipped exon/alternative 5' splice site events in moesin/ezrin/radixin-like protein 1 (IVC). A stop/splice-region SNV in trehalose phosphatase was detected with PYR exposure, highlighting coordinated disruption of the trehalose metabolism pathway. Across treatments, 68, 83 and 95 protein-coding genes with allelic variation were identified for ALB, PYR and IVC, respectively, including genes involved in detoxification, oxidative stress, cytoskeletal remodeling and transcriptional regulation.

Conclusions: Our findings reveal complex, drug-specific regulatory mechanisms in A. simplex, encompassing transcriptional remodeling, alternative splicing and functional SNVs. Novel modulation of trehalose metabolism and cytoskeletal genes, alongside potential roles for ABC transporters and RNA-binding proteins, suggests diverse adaptive strategies underlying anthelmintic tolerance. This study represents the first integrated transcriptomic and variant-level analysis of Anisakis under drug pressure and provides new insights into molecular resistance mechanisms in marine nematodes, with implications for therapeutic innovation and monitoring strategies.

Background: The sand fly genus Trichophoromyia Barretto, 1962 is one of the most diverse inthe subfamily Phlebotominae. The taxonomy and systematics of this group is complex due toboth a high similarity among species and unclear relationships among other sand fly groupswithin the subtribe Psychodopygina Galati, 1995. Despite their great relevance as vectors of Leishmania spp., few studies have explored the usefulness of molecular markers in studyingthe diversity of this group.

Methods: Here, we evaluated the use of barcode sequences of the cytochrome coxidasesubunit I gene (COI) for identifying several Trichophoromyia spp., by inferring intra- andinterspecific genetic distances, in addition to performing a set of several single-locus speciesdelimitation approaches using discovery methods. Moreover, we employed a multilocusdataset of four independent molecular markers (COI , ITS2 , 28S and PARA) to infer thephylogenetic species tree, estimate divergence times and delimit species under a validationmodel.

Results: The phylogenetic inferences confirmed the paraphyly of Trichophoromyia and Nyssomy ia Barretto, 1962. Thus, two new genera, named Reburrus gen. nov. and Shawmyia gen. nov., were proposed to accommodate sand flyspecies that did not fit in the aforementioned groups. Additionally, a new subgenus wasproposed: Trichophoromyia (Dilermandomyia) subg. nov., containing most speciesof Trichophoromyia . A recent speciation history was also estimated, with most of the speciesstudied diversifying during the Pleistocene. However, our dataset was insufficient to fullyresolve relationships within Trichophoromyia (Dilermandomyia) subg. nov. Many speciesshowed paraphyletic patterns in the gene trees, and some could not be reliably identified anddelimited using both COI barcodes and multilocus tools.

Conclusions: The sand fly genus Trichophoromyia exhibits a complex diversification history.Our phylogenetic inference and morphological observations of Nyssomyia and Trichophoromy ia, allowed us to propose new groups for the Psychodopygina subtribe. However, theprevalence of species-level paraphyletic patterns for Trichophoromyia (Dilermandomyia) subg.nov., showed that further assessment of this group requires a broader locus sampling combined with detailed morphological analysis.

Background: Spain's recent abrupt rise in numbers of registered pet dogs and cats has intensified the need for proper animal health care, as 60% of infectious diseases are zoonotic. While pathogen detection has improved through advances in molecular techniques, pet owners often fail to adhere to veterinary guidelines, increasing infection risks. Among the diagnostic tools available, faecal analysis plays a key role in detecting zoonotic parasites such as Giardia duodenalis, Cryptosporidium spp. and Toxocara spp. This study was designed to assess intestinal parasite prevalence in dogs and cats along with epidemiological trends.

Methods: Between 2013 and 2023, a total of 15,899 faecal samples from dogs and cats submitted to a reference laboratory of parasitology in Madrid (Spain) were analysed using Mini-FLOTAC^®, merthiolate-iodine-formalin (MIF), Baermann-Wetzel (for lungworms and S. stercoralis when indicated) and direct immunofluorescence assays, with molecular confirmation by polymerase chain reaction (PCR) when required. Epidemiological variables were statistically analysed.

Results: Overall, 26% of dogs and 21.4% of cats tested positive for at least one parasite. Protozoan infections were more prevalent overall, particularly G. duodenalis in dogs (16.0%) and Cystoisospora spp. in cats (7.8%). In contrast, helminth infections such as T. cati (7.6%) were more frequent in cats than in dogs. Dogs were more commonly infected by protozoa than helminths, while cats showed a more balanced distribution between both groups. Co-infections occurred in 13.6% of positive samples, with G. duodenalis being frequently involved. Puppies and kittens were more susceptible to infection, likely owing to an immature immune system. In animals from shelters, infection rates were higher than in owned pets. Seasonal variations were clearly observed, such that G. duodenalis peaked in winter and helminths in autumn. Over time, the prevalence of G. duodenalis increased, while that of Cystoisospora spp. declined.

Conclusions: These findings highlight the importance of parasite control for purposes of both animal and public health, and emphasize a need for regular faecal testing, deworming and improving owner awareness of parasites. To minimize zoonotic risks and improve pet health management, we would recommend standardizing diagnostic procedures and designing suitable veterinary interventions.

Background: Amblyomma maculatum is a tick with a broad host range that is undergoing an expansion of its range within the USA. When feeding the predilection sites on the host are the head and ears and due to the long mouthparts, it can cause significant lesions that can lead to infection. It has also been implicated as a vector of Hepatozoon americanum, the causative agent of canine hepatozoonosis and a spotted fever group Rickettsia, Rickettsia parkeri.

Methods: Two randomized, blinded, negative controlled studies were conducted to determine whether treatment with afoxolaner (NexGard^®, Boehringer Ingelheim) or a combination of afoxolaner, moxidectin, and pyrantel (NexGard^® Plus, Boehringer Ingelheim) effectively treats and controls infestations of A. maculatum on dogs. For each study, ten healthy dogs were randomly assigned to each treatment group. In one study there were three treatment groups: an untreated control, NexGard^®-treated group, and NexGard^® Plus-treated group. The other study had an untreated control group and a NexGard^® Plus-treated group. Dogs were infested with approximately 50 unfed adult A. maculatum prior to treatment for evaluation of efficacy against existing infestations and then three times after treatment for evaluation of persistent efficacy. In each study the appropriate treatment groups were treated with either NexGard^® or NexGard^® Plus with afoxolaner targeted at 2.5 mg/kg, and ten control dogs were untreated. For evaluation of efficacy, live ticks were counted and removed from each dog at 72 h after treatment or subsequent infestations.

Results: NexGard^® and NexGard^® Plus were > 99% effective against established infestations of A. maculatum compared with the control group (P < 0.0001). NexGard^® and NexGard^® Plus were ≥ 92% effective against reinfestation with A. maculatum through Day 31 of the studies (P < 0.0001).

Conclusions: The results of these studies demonstrate that NexGard^® and NexGard^® Plus administered once at or near the minimum recommended dose of 2.5 mg/kg afoxolaner is effective for the treatment of existing A. maculatum infestations and for the control of infestations through Day 31.

Background: Cerebral malaria (CM) is the most serious and fatal neurological complication of Plasmodium falciparum infection, which can cause death or long-term neurological sequelae. Neuronal injury is a primary cause of these sequelae in patients with CM; however, the underlying mechanisms remain incompletely elucidated. Hemozoin (Hz), the metabolic byproduct of hemoglobin digested by Plasmodium parasites, is closely associated with the severity of CM. However, it is not clear whether Hz is a direct contributor to neuronal injury.

Methods: C57BL/6 J mice were infected with the Plasmodium berghei ANKA (PbA) strain to induce experimental cerebral malaria (ECM). Hz deposition and neuronal injury in ECM mice brain tissues were assessed using histopathological staining. In vitro, primary cortical neurons were stimulated with purified hemozoin (pHz). Neuronal morphology, pHz internalization, and injury severity were assessed via transmission electron microscopy (TEM), live-cell imaging, and lactate dehydrogenase (LDH) assays, respectively. Furthermore, Mito-Tracker and JC-1 probes were used to analyze mitochondrial content and membrane potential, respectively. ATP assay kits were used to quantify cellular energy metabolism levels, while reactive oxygen species (ROS)/neuronal nitric oxide synthase (nNOS) fluorescent probes were used to assess oxidative stress and inflammatory response. Neurotransmitter alterations were analyzed by measuring glutamate (Glu) levels.

Results: In the cerebral cortex of ECM mice, significant Hz deposition and reduced neuronal nuclei (NeuN) expression levels were observed. Immunofluorescence (IF) staining demonstrated that pHz adhered to primary neurons in vitro, causing reduced dendritic arborization, axon rupture, and plasma membrane disruption. TEM and live-cell imaging confirmed that pHz was internalized into the cytoplasm of neurons. Furthermore, pHz induced mitochondrial structural damage and reduced mitochondrial content. Concurrently, pHz triggered mitochondrial dysfunction, characterized by diminished mitochondrial membrane potential (MMP), reduced ATP levels, and elevated ROS. In addition, pHz upregulated intraneuronal nNOS activity and caused a decrease in neurotransmitter levels.

Conclusions: This study provided the first evidence to our knowledge that Hz directly adhered to neurons and underwent internalization into its cytoplasm, thereby leading to neuronal injury. These findings elucidate a potential mechanism underlying neuronal injury in ECM and inform the development of adjuvant therapies targeting Hz.

Background: Mosquito saliva plays a key role in arbovirus transmission and pathogenesis. It was shown that saliva contains several molecules that are essential for blood feeding. Recently, bacteria were also reported to be present in the saliva of Aedes albopictus and Anopheles mosquitoes. Nevertheless, information on the bacterial communities in Aedes and Culex saliva is still scarce.

Methods: This study isolated and identified culturable fungal and bacterial colonies from saliva harvested from Aedes aegypti (laboratory strain) and Culex pipiens (field-collected) mosquitoes. 16S metagenomic sequencing was performed to identify bacterial communities in saliva and mosquito organs. Furthermore, it was assessed how these microbial communities were affected upon blood feeding and upon oral treatment with antibiotics and an antifungal drug.

Results: The fungal species Penicillium crustosum was identified in mosquito saliva. Culturable bacteria detected in mosquito saliva included Serratia marcescens, Serratia nematodiphila, Enterobacter spp., and Klebsiella spp., which were previously identified as mosquito or insect endosymbionts in the midgut or other organs. Analysis with 16S metagenomics showed that bacterial communities in saliva were more diverse than those in the midgut. Blood feeding did not affect the fungal or bacterial load in mosquito saliva. Oral treatment of adult mosquitoes with antibiotics or an antifungal drug resulted in a significant reduction of bacteria or fungi present in the mosquito saliva. Notably, co-incubation of the mosquito-borne Semliki Forest virus with saliva from antibiotic- or antifungal-treated mosquitoes triggered a decrease in viral infection in human skin fibroblasts compared with nontreated saliva.

Conclusions: These findings indicate that bacteria and fungi can be present in mosquito saliva and provide a foundation for further exploration of the impact of salivary fungi and bacteria on both vector competence and arbovirus infection in the mammalian host.

Background: Deep amplicon sequencing of nematode internal transcribed spacer 2 (ITS2), also referred to as the "nemabiome," has been increasingly used in veterinary hosts to study gastrointestinal nematodes. While post-sequencing bioinformatic pipelines such as DADA2 and mothur have been optimized, most researchers typically use the DADA2 pipeline in R. For optimal performance, DADA2 needs parameter tuning, which is hard for novices.

Methods: In this study, we present an implementation of the DADA2 pipeline within QIIME2 for nemabiome analysis and compare its performance against the commonly used R-based DADA2 pipeline. To evaluate performance against samples with known composition, we generated simulated nemabiome datasets representing canine, ruminant, and equine nematode communities. We also tested the pipelines using publicly available datasets from ten veterinary host species. For both pipelines, we evaluated differences in amplified sequence variant (ASV) generation, taxonomic classification, and diversity metrics. We also tested different Idtaxa parameter settings within the R DADA2 pipeline (classification threshold and bootstrap iterations) to understand its effects on nemabiome outcomes.

Results: While both pipelines showed minor discrepancies in relative abundance estimates, with minimal parameter optimization, QIIME2 outputs were closer to ground truth in simulated datasets. QIIME2 using the scikit Bayes classifier produced fewer unclassified taxa and more consistent species-level identifications compared with R DADA2's Idtaxa, particularly in complex communities. Community-level differences in beta diversity were primarily driven by differences in taxonomic assignment. Parameter testing revealed that lower classification thresholds in R DADA2 reduced the number of unclassified taxa but increased the risk of misclassification, highlighting the need for careful parameter selection and reporting.

Conclusions: With minimal parameter tuning, QIIME2 outperformed the R pipeline in taxonomic resolution, and improved reproducibility by provenance tracking. Our findings emphasize how bioinformatics pipeline choices impact nemabiome outputs including the number of species detected, ranks of abundant taxa, and alpha and beta diversities. We provide a reproducible and user-friendly QIIME2 workflow suitable for researchers seeking standardized analyses of ITS2 nemabiome data.

Background: Aedes aegypti is the primary vector of arboviruses, including dengue, Zika and chikungunya, representing a major global public health concern. Owing to the lack of effective vaccines and specific therapeutic options for these infections, vector control remains the main strategy to limit their spread. Traditionally, vector control has relied on extensive use of insecticides combined with the elimination of breeding sites. However, in addition to selecting for insecticide-resistant mosquitoes, concerns have arisen about behavioural effects induced by insecticides, particularly repellency - defined as the ability of a chemical compound to trigger avoidance behaviour in insects, thereby reducing their exposure to treated surfaces. This systematic review aimed to synthesise current knowledge on repellent effects of certain insecticides on A. aegypti.

Methods: A literature search was conducted in the databases Virtual Health Library (BVS), PubMed® and Scientific Electronic Library Online (SciELO), following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. A total of 46 original studies published between 1990 and 2023 were included.

Results: Altogether, 433 bioassays were analysed, of which 69.8% reported repellent effects. The most common methods used to assess repellency were excito-repellency chambers, HITSS assays and the arm-in-cage test. Pyrethroids were used in 86.6% of repellency assays, followed by organochlorines (9.4%). Regarding the resistance profile of tested mosquito populations, susceptible populations exhibited higher frequencies of contact (92.2%) and spatial (77.3%) repellency behaviours than resistant ones (74.1% and 44.0%, respectively).

Conclusions: Our findings indicate that insecticide-induced repellency is common and may interfere with the effectiveness of chemical control strategies. Nevertheless, studies addressing the underlying molecular and sensory mechanisms involved in repellent perception remain scarce.