Terebelliform annelids can serve as intermediate hosts for blood flukes of marine fishes and sea turtles. The scarcity of global reports on the life cycles of these parasites reflects the need for further exploration. Our objective was to further our knowledge of the diversity and life cycles of blood flukes in coastal South Carolina (SC), United States. Four species of terebellids ( Enoplobranchus sanguineus , Amphitrite ornata , Pista palmata , and Streblosoma hartmanae ) were collected (total N = 201) at low tide from September 2025 to February 2026 at two localities in SC. Twelve specimens of three terebellid species were infected, and parasites were identified by sequencing the LSU and ITS2 regions of ribosomal RNA and the COI mitochondrial DNA gene. Enoplobranchus sanguineus (12.9% prevalence, 4/31) and A. ornata (3.6% prevalence, 2/55) were infected with the previously reported blood flukes Neospirorchis sp. Neogen14 (Unicaecidae) and Cardicola parvus (Aporocotylidae), respectively. No specimens of S. hartmanae were infected (0/86). One new aporocotylid with mature sporocysts containing microcercous cercariae was discovered in P. palmata (20.7% prevalence, 6/29). Phylogenetic analysis revealed that sequences of the new aporocotylid were in a well-supported clade with sequences of Braya spp. and Cardicola abu . The fish host of this parasite has yet to be discovered to fully elucidate the life cycle of this new aporocotylid.
Microsporidia are obligate intracellular parasites with a wide host range. To elucidate the relationship between Enterocytozoon bieneusi infection dynamics and evolution within China’s sika deer population, 466 fresh fecal samples were collected from May to October 2024 across four provinces, Jilin, Liaoning, Heilongjiang, and Shandong. The ITS region of ribosomal RNA (rRNA) was amplified using nested PCR, enabling the identification and genotyping of E. bieneusi. The overall prevalence of E. bieneusi in sika deer was 44.4% (207/466). Province-specific prevalence was highest in Shandong (76.0%, 38/50) and Heilongjiang (75.3%, 67/89), followed by Jilin (38.7%, 65/168), and lowest in Liaoning (23.3%, 37/159). The prevalence in autumn (44.9%) was slightly higher than in summer (41.7%), and the prevalence in adult deer (48.7%) was found to be significantly higher than in young deer (31.6%). In this study, we identified a total of 11 E. bieneusi genotypes, including 9 known genotypes (BEB6, HND-II, LND-I, JLD-XIX, CHN-F1, HLJD-I, HLJD-III, Type IV, and COS-I) and 2 novel genotypes (named JL-D-4 and JL-D-5). Among them, BEB6 was the dominant genotype (38.7%, 80/207). This study enriches the genotype data of E. bieneusi , reveals the prevalence of E. bieneusi in some sika deer populations, and provides a reference for subsequent epidemiologic research as well as prevention and control efforts.
Background : Zoonotic cutaneous leishmaniasis (ZCL) caused by Leishmania major remains a major public health concern in Morocco, especially in certain semi-arid and pre-Saharan regions where ecological conditions favor transmission. In recent decades, the epidemiology of ZCL has evolved significantly, particularly in its temporal patterns and the persistence of transmission in major endemic provinces. In this study, we adopted a retrospective observational approach to explore the long-term spatiotemporal dynamics of ZCL across nine endemic provinces in Morocco and to assess its potential environmental, socio-economic, and demographic determinants using a descriptive analytical framework. Principal findings : A total of 58,880 ZCL cases were reported during the study period between 1995 and 2022, with incidence peaks observed in 2010 and 2018, reflecting cyclical epidemic dynamics. The Drâa-Tafilalet region accounted for the highest disease burden, with Zagora province identified as the main epidemiological hotspot. Children aged ≤10 represented the most affected age group (40.8% of total cases), whereas infection rates did not differ significantly between the sexes (female-to-male ratio = 1.17). Higher incidence was observed in hyper-arid provinces characterized by sandy-clay-loam soils. However, incidence was not statistically correlated with either environmental or social variables ( p > 0.05). Conclusions : ZCL remains endemic in Morocco, with persistent hotspots linked to specific ecological contexts. These findings highlight the importance of geographically targeted surveillance and control measures to support the integration of environmental indicators into risk monitoring and early-warning systems for leishmaniasis.
The European brown hare (Lepus europaeus) is an ecologically important small game mammal whose populations have declined in many European countries. A total of 370 faecal samples collected between 2022 and 2024 from hares in the Czechia and Slovakia were molecularly screened for the presence of Cryptosporidium spp. (18S rRNA, gp60), Giardia intestinalis (TPI), Encephalitozoon spp., and Enterocytozoon bieneusi (ITS). qPCR analysis confirmed the presence of Cryptosporidium spp. (n = 15; 4.1%; CI: 2.5-6.6%), Encephalitozoon spp. (n = 34; 9.2%; CI: 6.7-12.6%), and E. bieneusi (n = 16; 4.3%; CI: 2.6-6.9%). Sequence analyses revealed the presence of C. parvum (n = 2), C. sciurinum (n = 3), C. ubiquitum (n = 5), and Cryptosporidium sp. deer mouse genotype IV (n = 5). At the gp60 locus, C. parvum belonged to subtypes IIaA15G1R1 and IIaA23G1R1, C. sciurinum to subtypes VIIIcA10G2R1 (n = 2) and VIIIbA11G1R1 (n = 1), and C. ubiquitum to subtypes XIIa (n = 3) and XIId (n = 2). Five E. bieneusi genotypes were identified: D (n = 8), CHALT1 (n = 1), WildBoar3 (n = 1), CHN-F1 (n = 2), and C (n = 4). Encephalitozoon sequencing revealed E. cuniculi genotypes I (n = 6), II (n = 26), and III (n = 1), and E. hellem (n = 1). Giardia intestinalis was not detected. Cryptosporidium sp. deer mouse genotype IV is reported here for the first time in Europe.
Scabies is a common health concern, particularly among infants and children from socioeconomically disadvantaged households. This single-center retrospective observational cohort study describes scabies phenotypes, associated factors, treatment patterns, complications, and 12-month scabies-related re-attendance among diagnosed children between March 2018 and March 2023. Data from 1,094 children living in Turkey were extracted from medical records. Patients were classified as having classic, crusted, or nodular scabies for descriptive analyses; crusted and nodular scabies were combined as variant scabies only for selected regression models. Logistic regression was used to identify factors associated with variant scabies and hospitalization. Most cases occurred in fall and winter (67.6%). Crusted and nodular scabies were identified in 93 (8.5%) and 18 (1.6%) patients, respectively. Underlying abnormalities were reviewed separately for crusted and nodular scabies. Among hospitalized children, 61.5% had variant scabies, and clinically suspected secondary bacterial infection was documented in 77 of 78 hospitalized patients. Cultures were obtained from all hospitalized children; among culture-positive cases, Staphylococcus aureus was the most frequently isolated organism. Among children who re-attended within one year, permethrin was the most frequently documented treatment (92.3% of documented treatments). Underlying disease and younger age were independently associated with variant scabies, whereas underlying disease, variant scabies, and younger age were independently associated with hospitalization. Re-attendance was more frequent in variant scabies ( p < 0.001). Crusted scabies warrants careful evaluation for underlying abnormalities and secondary infection, while nodular scabies should be interpreted as a distinct hypersensitivity-related phenotype. These findings support proactive household-level management and follow-up.
BACKGROUND:Soil-transmitted helminths (STHs) and other enteric parasites remain endemic in rural communities of northwestern Argentina. Environmental surveillance may provide additional information on transmission dynamics and community exposure. RESULTS:We analyzed 116 soil samples collected from 28 sites, including 27 households in Solazuti and one rural school in El Cedral (Orán Department, Salta Province, Argentina), using multi-parallel real-time quantitative PCR targeting intestinal protozoa, STHs, zoonotic nematodes, and Acanthamoeba spp. Parasites were detected in outdoor built environments, with 44% of Solazuti households showing at least one positive sample. Ascaris lumbricoides and Toxocara canis were the most frequently detected helminths and showed distinct spatial patterns. Detection of A. lumbricoides was strongly associated with the presence of other parasites (OR 18.11, 95% CI 8.30-38.6; p < 0.0001). Additionally, A. lumbricoides-positive households harbored significantly more parasite species than A. lumbricoides-negative households (p = 0.0343), and A. lumbricoides frequency correlated with parasite richness (Spearman r = 0.4145, p = 0.0316). Acanthamoeba spp. was nearly ubiquitous (96.4% of sites), including the school environment, with a 75% contamination rate. CONCLUSIONS:Molecular analysis of soil samples revealed spatially heterogeneous but sustained exposure to multiple pathogens in rural outdoor environments. Soil-based qPCR may complement traditional surveillance by providing community-level data independent of human sampling.
This study aimed to assess the prevalence and risk factors of molecular markers of Plasmodium (P.) falciparum resistance to sulfadoxine-pyrimethamine (SP) among pregnant women in Bobo-Dioulasso, and to examine their association with maternal anaemia. This cross-sectional study was conducted between October and December 2022 among 288 SP-naïve pregnant women attending antenatal care at the Centre Médical Urbain of Lafiabougou. Dried blood spots were collected, and P. falciparum infection was detected using quantitative polymerase chain reaction (qPCR) targeting the varATS gene. Positive samples were genotyped for mutations in the Pfdhfr and Pfdhps genes using a nested PCR approach followed by restriction fragment length polymorphism analysis. Multivariable logistic regression models were used to identify predictors of resistance markers and maternal anaemia. Among the 172 qPCR-confirmed P. falciparum–positive samples, the Pfdhfr triple mutant haplotype (N51I, C59R, S108N) was detected in 33.7% of cases, while the Pfdhps A437G mutation was present in 82.7%. No Pfdhfr I164L or Pfdhps K540E mutations were observed. Gestational age was independently associated with carriage of the triple Pfdhfr mutation (adjusted OR = 2.2, 95% CI: 1.1–4.5). Both gestational age (adjusted OR = 2.5, 95% CI: 1.2–4.9) and infection with parasites carrying the triple Pfdhfr mutation (adjusted OR = 3.9, 95% CI: 1.8–8.3) were significant predictors of maternal anaemia. The relatively high prevalence of SP resistance markers indicates sustained drug pressure in this setting. Although SP appears to remain effective for intermittent preventive treatment in pregnancy, continued molecular surveillance is warranted to inform malaria control policies.
While various haemoparasites have been reported from anuran hosts, this study provides the first published record of dactylosomatid parasites (Dactylosomatidae Jakowska & Nigrelli, 1955 emend. Levine, 1971) in frogs from Slovakia. Of the 239 anurans screened for apicomplexans, 67 individuals belonging to three species of water frogs, Pelophylax esculentus (Linnaeus, 1758), P. ridibundus (Pallas, 1771), and P. lessonae (Camerano, 1882) were found to be infected with haemogregarines of the genus Dactylosoma Labbé, 1894. Our results demonstrate that the haemogregarine found in all three species of water frogs, collected from three different localities in western Slovakia, represents a single taxon that morphologically resembles the type species, Dactylosoma ranarum (Kruse, 1890). Based on comprehensive morphometric, morphological, and molecular data from newly collected isolates from Pelophylax frogs, including the type host, we provide an integrative assessment of Dactylosoma cf. ranarum. We also conducted molecular screening of dipterans collected from the study sites, which could serve as potential vectors of the parasite, but no representatives of Dactylosoma were detected in any of the examined specimens.
The mitochondrial genome of kinetoplastids, organized in a unique DNA net called the kinetoplast, presents significant complexity compared to other eukaryotic mitochondrial genomes. Minicircles, which constitute over 90% of the kinetoplast mass, are essential for the post-transcriptional editing of maxicircle transcripts. This study focuses on Trypanosoma lainsoni, a trypanosomatid first discovered in northern Brazil and later reported in Argentina. Utilizing a combined second and third generation sequencing approach, we conducted an in-depth analysis of the kDNA minicircles of three Argentinian isolates of T. lainsoni. Through de novo assembly, minicircle molecules with two conserved regions interspaced by two hypervariable regions 180° apart, were identified. Guide RNAs encoded within hypervariable regions were inferred and the mitochondrial mRNA editing cascades mediated by these guide RNAs were fully reconstructed. We also developed a PCR-based approach for the detection and confirmation of T. lainsoni kDNA using minicircle-specific primers. A deeper analysis on the diversity of minicircle hypervariable regions revealed that the three isolates correspond to different genotypes circulating in the region. This research significantly advances our understanding of the genomic architecture of T. lainsoni and offers valuable tools for its molecular identification, differentiation from other trypanosomes, and the study of kinetoplastid biology.
Thelazia callipaeda, commonly known as the oriental eyeworm, is a vector-borne parasitic nematode that infects the ocular tissues of a wide range of mammalian hosts, including dogs, cats, wildlife, and humans. Historically confined to East and Southeast Asia, T. callipaeda has emerged over the past 2 decades as a significant zoonotic parasite in Europe, with an expanding geographic distribution driven by the spread of lachryphagous drosophilid fruit fly vectors of the genus Phortica. This review synthesizes current knowledge on the taxonomy, epidemiology, biology, genetic diversity, pathogenesis, and control of T. callipaeda, with particular emphasis on its One Health relevance. Molecular studies reveal low but structured genetic variability, characterised by a single predominant haplotype circulating in Europe and high haplotype diversity in Asian populations, reflecting long-term endemicity and distinct transmission dynamics. Clinically, infection can result in ocular irritation ranging from mild conjunctivitis to severe keratitis and corneal ulceration, with dogs acting as the primary domestic reservoir and wildlife sustaining sylvatic transmission cycles. Human infections, though underreported, are increasingly recognised and pose a growing public health concern. Effective management relies on mechanical worm removal, macrocyclic lactone treatment and prevention, and integrated surveillance of animal hosts and vectors. Given the influence of climate change, animal mobility, and environmental factors on vector ecology, coordinated One Health strategies are essential to mitigate the continued spread and zoonotic impact of this emerging eyeworm.
Ectoparasiticides are essential for preventing insect bites and transmission of arthropod-borne pathogens such as Leishmania spp. Traditionally, their anti-feeding efficacy is evaluated in vivo using sedated dogs exposed to sand flies. To comply with 3R principles (replacement, reduction and refinement) and to avoid animal exposure, this study aimed to develop an ex vivo feeding model to assess the efficacy of a dinotefuran, permethrin, and pyriproxyfen (DPP) [Vectra®3D] combination against Phlebotomus perniciosus as an alternative to in vivo testing. Twelve dogs were assigned to either a DPP-treated group (n = 6) or an untreated control group (n = 6). On days -7, 1, 7, 14, 21, and 28, dogs were sedated and exposed for one hour to 50 (±5) female sand flies. In parallel, hair collected from each dog was used in an ex vivo artificial feeding system. After exposure, sand flies were assessed for feeding and survival rates. Anti-feeding efficacy was the primary criterion, while insecticidal efficacy was evaluated as a secondary outcome. In both models, control groups showed feeding and survival rates above 31% and 90%, confirming a reliable challenge. The treated group showed significantly lower feeding and survival rates than the control group (p < 0.05). In both models, DPP demonstrated a strong anti-feeding effect in dogs for one month, with efficacy above 80%. This first direct comparison of in vivo and ex vivo models confirms DPP’s fast and lasting protection against Ph. perniciosus and positions the ex vivo model as a promising 3R-aligned alternative for studying vector-borne disease prevention.
Effective surveillance of invasive Aedes mosquitoes, particularly Ae. aegypti and Ae. albopictus, is essential for early detection, risk assessment, and optimization of control strategies such as the Sterile Insect Technique (SIT). Although BG-Sentinel 2 traps are widely used in Europe, their performance under eastern Mediterranean conditions remains insufficiently evaluated. We assessed the attractiveness of BG-Sentinel 2 traps baited with BG-Lure, dry ice (CO2), their combination, and unbaited controls using a Latin square field design in Larnaca and Nicosia, Cyprus. A total of 1,649 mosquitoes were collected. In Larnaca, Ae. aegypti occurred at low densities. Male captures were consistently low and unaffected by attractants, while female captures increased significantly only when BG-Lure and CO2 were combined (RR = 2.7, 95% CI: 1.6–4.5). In Nicosia, Ae. albopictus was abundant, but responses were sex-specific. Male captures were significantly reduced by CO2 alone (RR = 0.297, 95% CI: 0.175–0.467), with a weaker reduction when combined with BG-Lure, while female captures were not significantly affected by any treatment. For Culex pipiens, CO2 strongly increased female captures (RR ≈ 16), irrespective of BG-Lure, whereas males showed only a modest response to BG-Lure alone. Overall, attractant efficacy was highly species- and sex-specific. Standard baiting strategies did not consistently enhance detection of invasive Aedes mosquitoes and, in some cases, reduced male captures, with implications for SIT programs. These findings emphasize the need for local validation of surveillance protocols particularly at points of entry.
Zoonotic cutaneous leishmaniasis (ZCL) caused by Leishmania (L.) major is the most widespread form of CL worldwide, with hundreds of thousands of cases annually. It is associated with significant morbidity and economic burden, particularly in North Africa and the Middle East. For over a century, the zoonotic transmission of ZCL, involving wild gerbils (Muridae: Gerbillinae) as the reservoir and Phlebotomus (Ph.) papatasi as the vector, has been considered an established fact, with no contradictory debate. However, the pronounced endophilic and anthropophilic behaviors of Ph. papatasi, the exposure of multiple and large lesions, although with relatively limited persistence, to sandflies during their seasonal activity, and the simultaneous detection of human and L. major DNA within the vector, suggest a contribution of humans to the parasite cycle. Conversely, the high incidence of the disease in anthropized foci and the high prevalence of family cases highlight the role of humans in amplifying transmission. The experiments carried out by Adler and Theodor in the 1920s in Palestine and the more recent studies done by Fatemi et al. (2018) in Iran that identified L. major infectious metacyclic forms in Ph. papatasi fed on ZCL lesions reinforce this hypothesis. Additional xenodiagnoses-based approaches could provide definitive answers and lead to better management and control of ZCL, leading to a recommendation of updated preventive measures at the household level. This review aims to report on what is currently known about the zoonotic transmission of L. major, and to develop arguments supporting the hypothesis that humans play a potential role as "reservoir host" of the parasite, albeit a complementary one.
Trichostrongylus spp., zoonotic soil-transmitted nematodes, affect both livestock and humans globally. In Thailand, human trichostrongylosis has been reported, but no systematic study examining livestock and humans in the same locality has been conducted. This study aimed to address this gap in Satun Province, southern Thailand by characterizing the disease's epidemiologic status among livestock, farmers, and the environment using both microscopic and molecular techniques. Human risk factors were assessed via questionnaires. Stool samples were collected from livestock farmers, their families, and herbivores in Nongkhai, Ratchaburi, and Satun Provinces. Vegetable samples were also obtained from households and local markets. Human and livestock feces were examined using the modified Kato-Katz method and a Mini Parasep® solvent-free fecal parasite concentrator. Total DNA was then extracted from fecal and vegetable sediments, and a 211 bp fragment of the ribosomal internal transcribed spacer 2 (ITS2) gene was amplified using a polymerase chain reaction (PCR). PCR products were digested with HinfI and analyzed via electrophoresis to identify Trichostrongylus species. In Satun, Trichostrongylus colubriformis monoinfection was found in 12 of 221 (5.4%) residents, while co-infection with T. colubriformis and T. axei occurred in 3 (1.4%). Herbivore feces and vegetable sediments from Satun also tested positive. No human cases were found in Nongkhai or Ratchaburi. Risk factors included education level, occupation, and outdoor toilet use. Most infected individuals in Satun were asymptomatic. Despite this, public health interventions should be implemented, targeting the human-livestock-environment interface to control the disease effectively.
The long-term seasonal parasitism patterns, hereafter referred to as stylopization, and within-nest developmental stage distributions of Vespa-associated Xenidae (Strepsiptera) remain poorly characterized in South Korea. We compiled 15 years of data (2008-2023) based on trapped Vespa individuals from multiple regions and quantified stylopization rates for two Xenidae species, Xenos oxyodontes and Xenos moutoni, based on diagnostic characters visible on stylopized hosts. Seasonal occurrence was summarized primarily from female Xenos specimens, which can be identified to species, and host-association patterns were assessed where possible. To complement the trap-based records, we dissected 22 Vespa analis colonies collected in September 2020 and documented strepsipteran developmental stages within the nests. Across 39,610 examined host individuals, stylopization rates and seasonal occurrence differed between the two Xenos species and among host taxa, indicating distinct host-use and temporal activity patterns. Importantly, the nest dissections provided direct observations of late-season developmental stage compositions, enabling us to compare within-nest reproductive/developmental stages to those inferred from trap-detected stylopized hosts. This comparison revealed that key reproductive-stage information may not be captured by trap records alone. Thus, integrating long-term trap data with nest-level observations can clarify species-specific phenology and developmental/reproductive stage distributions in Vespa-associated Xenidae, providing a better empirical context for interpreting stylopization intensity and developmental timing in field populations.
Microsporidia are obligate intracellular eukaryotes infecting a wide range of hosts and cell types. The development of in vitro culture models is essential for studying host–parasite interactions and the pathogenesis of microsporidian infections. In this study, we compared the infection efficiency of three human pathogenic species of the genus Encephalitozoon (Encephalitozoon intestinalis, E. hellem, and E. cuniculi) across six cell lines: TC7, HT-29, HCT 116, T84, Vero, and MRC-5. Infection rates and the surface area of parasitic foci were determined after labeling microsporidia by fluorescence in situ hybridization (FISH). Both parameters varied significantly according to the cell line and Encephalitozoon species. TC7 cells consistently supported the highest infection rates, whereas HCT 116 cells were the least permissive. The surface areas of foci were primarily species-dependent, with larger foci observed for E. cuniculi and smaller ones for E. intestinalis. In conclusion, these results revealed marked differences in invasive and proliferative dynamics depending on the Encephalitozoon species and cell line and highlight the crucial impact of cell line selection on in vitro experimental outcomes. This work provides a foundation for improving and standardizing experimental models for future experiments on Encephalitozoon spp.
In this study, adults and plerocercoids of S. mansoni were cultivated in vitro to systematically analyze the components of the excretory–secretory proteins (ESPs) of Spirometra mansoni. Afterwards, the differentially expressed proteins (DEPs) were identified and protein components were examined using the Data Independent Acquisition (DIA) mode. A total of 944 proteins were identified, including 580 plerocercoid-specific proteins, whereas no specific proteins were found in adults. Quantitative analysis revealed that 607 proteins were significantly differentially expressed, with 390 upregulated in the plerocercoid group, and 217 upregulated in the adult group. Gene Ontology functional annotation revealed that the upregulated proteins in the plerocercoid group were significantly enriched in functions such as nitrogen compound metabolism, proteasome core complexes, and ion binding. Kyoto Encyclopedia of Genes and Genomes pathway enrichment revealed that the DEPs were strongly correlated with signal transduction, signal transportation, and catabolism pathways. Moreover, metabolic network analysis revealed that key pathways included the pentose phosphate pathway and glycolysis/gluconeogenesis. In addition, indirect ELISA revealed that immunization of mice with ESPs induced a Th1/Th2 mixed immune response, dominated by a Th1 response. Cytokine detection further verified that ESPs had good immunogenicity, and could activate both humoral and cellular immune responses. This study revealed, for the first time, the differential expression profile of ESPs between adults and plerocercoids of S. mansoni. These findings offer a potential reference for the diagnosis and prevention of sparganosis.
Granulocytic anaplasmosis is a zoonotic disease that affects various domestic mammals (dogs, horses, and, more rarely, cats). In ruminants, it is better known as tick-borne fever (TBF) and is responsible for significant economic losses on European livestock farms, mainly due to a drop in milk production, abortions, and immunosuppression, which can lead to secondary infections. The disease is caused by the strictly intracellular bacterium Anaplasma phagocytophilum, whose biological vectors are ticks of the genus Ixodes. Other blood-feeding arthropods may be involved in transmitting this bacterium, notably Stomoxys calcitrans, a major ectoparasite of livestock that is implicated in transmitting other pathogens, including bacteria of the genus Anaplasma. This study aimed to evaluate the potential of S. calcitrans to act as a mechanical vector of A. phagocytophilum under laboratory conditions. Two experimental models were employed: one mimicking immediate transmission, and the other delayed transmission. In both models, A. phagocytophilum DNA and RNA were detected in S. calcitrans for the first time, but no traces of the bacterium’s DNA or RNA were found in the glass feeder’s blood. Further research is needed to confirm these findings through field studies investigating the presence of the bacterium in flies under natural conditions. This study also describes two original infection models of stable flies designed to reproduce their ex vivo blood-feeding, promoting alternative experimental approaches in accordance with animal welfare regulations and 4R principles.
Previous studies have shown that recombinant Trichinella spiralis serine proteinase (rTsSPc) promoted larval invasion of the gut epithelium, but its regulatory role on macrophage polarization is not clear. Immunofluorescence assay (IFA) confirmed specific rTsSPc’s binding to RAW264.7 macrophages. The results of qPCR, Western blot, ELISA, and flow cytometry showed that rTsSPc significantly upregulated M1 macrophage markers (iNOS and CD86) and pro-inflammatory cytokines (TNF-α and IL-6), but not M2 markers (Arg1, CD206) and anti-inflammatory cytokines (IL-10, TGF-β). However, intestinal infective larvae (IIL) excretory-secretory antigens (ESAs) induced M2 polarization. Western blot revealed that rTsSPc activated the classical NF-κB pathway, as evidenced by increased phosphorylation levels of IKKβ, IκB-α, and NF-κB p65. Pretreatment of macrophages with NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC) effectively suppressed rTsSPc-induced M1 polarization, decreased pro-inflammatory cytokine secretion, and reduced nitric oxide (NO) production. Functionally, rTsSPc-induced M1 polarization significantly enhanced antibody-dependent cell-mediated cytotoxicity (ADCC) of macrophages killing newborn larvae, but PDTC pretreatment resulted in a 41.62% reduction in cytotoxicity. Our results showed that rTsSPc bound specifically to macrophages and induced M1 polarization by activating the classical NF-κB pathway, thereby enhancing macrophage-mediated ADCC killing of newborn larvae. The findings indicated that TsSPc might strengthen host protective immunity via ADCC killing of larvae, and TsSPc may be considered a potential candidate antigen for developing anti-Trichinella vaccines.
The DBLMSP gene family in Plasmodium falciparum encodes surface antigens involved in immune evasion and red blood cell invasion, yet its extensive polymorphism has long defied straightforward classification. While analyzing DBLMSP1 sequences from samples collected along the China–Myanmar border, we found that haplotypes could not be readily explained by standard population genetic models. Instead, comparative alignment and BLAST analysis revealed that DBLMSP1 and DBLMSP2 consist of discrete, recombinable sequence modules, flanked by conserved upstream and downstream regions. This led us to propose a modular framework that redefines allele structure as combinations of well-defined building blocks with consistent boundaries and positional constraints. Through global mining of DBLMSP sequences, we identified nine genotypes each for DBLMSP1 and DBLMSP2, with modules labeled sequentially (e.g., 1M2a, 1M3c, and 2M3b). Some modules were shared across paralogs, notably the identical sequence of DBLMSP1 module 1M3c and DBLMSP2 module 2M3a, suggesting historical inter-locus recombination. In the dominant genotype DBLMSP1-1, nucleotide diversity and Tajima’s D peaked within variable modules, whereas conserved structural elements, including the receptor-binding cleft and SPAM domain, were under purifying selection. Patterns of long-range linkage disequilibrium aligned with module junctions, suggesting that modular structure may shape recombination patterns independently of selection. Modular recombination has been widely recognized in viral systems and multigene families such as var, but its relevance in DBLMSPs has been underappreciated. By applying this framework to P. falciparum DBLMSPs, we aim to provide a useful perspective for understanding their structural diversity and evolutionary dynamics, with implications for immunogen design and parasite surveillance.
