Infective Larvae Research Articles

Experimental arboviral infection of mosquito larvae: A novel approach for vector competence studies

Vector competence studies in mosquitoes present valuable opportunities to explore arboviral transmission and virus-vector interactions. However, oral infection studies in mosquitoes can be challenging. An alternative approach is to infect mosquitoes during their aquatic larval stage, resulting in the emergence of infected adults. To investigate the potential of this method, Culex pipiens biotype molestus larvae were infected with Usutu virus (USUV, Orthoflavivirus usutuense). For this purpose, larvae were exposed to USUV-infected mammalian and mosquito cell cultures for 24 h before being reared to adults. Subsequent analysis via RT-qPCR revealed that the Culex larvae successfully acquired USUV from the infected cells and exhibited high susceptibility to infection. Immediately after emergence, 32.10 % (26/81) of male and 41.03 % (16/39) of female mosquitoes tested positive for USUV RNA. Notably, females that were incubated for 15 days post-emergence demonstrated even higher infection rates, reaching 100.00 % (23/23). In addition, viral RNA and infectious particles were detected in some saliva samples, indicating the potential for transmission. This experimental infection of mosquito larvae thus offers the opportunity to produce infected adult mosquitoes for studies enhancing our understanding of virus-vector interactions, co-infections, and transmission routes. Such research contributes to better public health strategies addressing arboviral diseases.

Tetracycline and Oxacillin Act Synergistically on Biofilms and Display Increased Efficacy In Vivo Against Staphylococcus aureus.

Oxacillin (bactericidal) and tetracycline (bacteriostatic) are clinically relevant antibiotics that are routinely prescribed to treat Staphylococcus aureus infections but not conventionally used in combination. There is an urgent need for treatment regimens that can act upon biofilms during infection, associated with chronic infections on indwelling devices, as well as acute planktonic (systemic) infection. Here we show that in an in vitro model oxacillin and tetracycline act synergistically against S. aureus UAMS-1 biofilms, reducing the concentration of both antibiotics necessary to eradicatean established biofilm. Using an in vivo zebrafish larval infection model with S. aureus NewHG, they display improved bacterial clearance compared to each drug alone and can counteract a loss of host phagocytes, an important innate defence against S. aureus. In these cases, the bacteriostatic nature of tetracycline enhances rather than dampens the bactericidal action of oxacillin, although an exact mechanism remains to be elucidated. We suggest a dual therapy could be of clinical use against biofilm-forming S. aureus and has a potential use in patients with a compromised immune system.

Tenebrio molitor (Coleoptera: Tenebrionidae) as an alternative host for the study of pathogenicity in Candida auris

BackgroundCandida auris, a multidrug-resistant fungal pathogen, has emerged as a significant global health threat due to its high transmission and mortality rates, especially in healthcare settings. ObjectiveThis study aimed to establish the larvae of the coleopteran Tenebrio molitor (mealworm) as an in vivo model to evaluate the virulence of different C. auris strains. MethodsT. molitor larvae were inoculated with varying doses and strains of C. auris. Mortality rates were monitored, melanization responses, and phenoloxidase activity were assessed. Histopathological analyses were conducted to observe tissue invasion by the yeast cells. Additionally, a biofilm formation test was included as a complementary analysis to determine if biofilm production would influence the virulence of the C. auris strains. ResultsA dose-dependent increase in mortality was observed, with the highest fungal load leading to the highest mortality rates. The study also revealed significant differences in virulence among the strains, with those from Kuwait and the reference strain CBS 10913 showing the highest pathogenicity. Melanization rates were significantly higher in infected larvae, indicating an active immune response. The histopathological analysis revealed the presence of C. auris cells within the tissue of T. molitor larvae. However, the biofilm formation complementary test did not show a significant difference in virulence among the different clades of C. auris. ConclusionThe T. molitor model effectively demonstrated the pathogenic potential and virulence differences of C. auris strains. Strains from Kuwait and the reference strain CBS 10913 exhibited the highest virulence, causing 100% mortality within 24 hours. The model also highlighted significant biofilm formation and melanization responses, correlating with fungal burden. This insect model provides a valuable and cost-effective tool for preliminary virulence screening of clinical yeast strains, offering insights into host-pathogen interactions and the potential for evaluating antifungal treatments in vivo.

The role and mechanism of BmsPLA2-1-1 in the IMD pathway in silkworm, Bomybx mori

Lepidoptera are a major source of pests in agriculture and forestry, investigating the immune mechanisms of their model species, Bombyx mori, can provide valuable insights into improving pest management. Although the phospholipase A2 (PLA2) and immune deficiency (IMD) pathway have been extensively investigated, their relationship remains unclear. Here, we found that bacterial infection of silkworm larvae significantly upregulated BmsPLA2-1-1 expression and knockdown of BmRelish in the IMD pathway suppressed this response. Likewise, reducing BmsPLA2-1-1 expression significantly downregulated IMD pathway-related genes, including BmRelish, BmImd, and BmPGRP. In contrast, overexpression of BmsPLA2-1-1 significantly upregulated BmRelish and BmImd expression, suggesting a functional crosstalk between BmsPLA2-1-1 and the IMD pathway in silkworms. Additionally, BmPLA2-1-1 interacted with BmHsp60, BmCNBP, BmCfp1, and BmPFD3. Reducing BmRelish resulted in decreased expression of BmHsp60A, BmCfp1, and BmPFD3, but not BmCNBP, in infected larvae. Overexpression BmHsp60A, BmCfp1, or BmPFD3 led to a significant upregulation of BmRelish and BmImd expression. These suggest that BmHsp60A, BmCfp1, and BmPFD3 are involved in functional crosstalk between BmsPLA2-1-1 and the IMD pathway in silkworms. These findings demonstrate the functional crosstalk and mechanism between BmsPLA2-1-1 and the IMD pathway, revealing a new mechanism in the insect immune network.

Temporal increase in ticks and pathogen prevalence in the small mammal part of the Lyme disease cycle in northern Europe

AbstractLyme disease is an emerging infectious disease and the most common vector‐borne zoonosis in the northern hemisphere. The pathogen that causes Lyme disease in Europe is vectored by the generalist tick Ixodes ricinus, and the emergence of Lyme disease is partly linked to how climate warming affects tick distribution and abundance. However, we lack long‐term data on tick infestations and infection prevalence in the main hosts involved in the transmission cycle. Here, we quantified the temporal trends (2014–2022) of I. ricinus infestations and the prevalence of Borrelia burgdorferi sensu lato in small mammalian hosts and linked annual variation to host abundance and climate in Norway. We found that tick infestations for both larvae (21% per year [95% CI 18–25]) and nymphs (18% [11–26]), and infection prevalence (14% [8–20]) increased over the period and were negatively associated with rodent abundance. Additionally, warmer years were associated with increased larval tick infestations on hosts. The combination of a temporal increase in both larval tick infestation and infection prevalence in hosts likely results in increased production of infected nymphs. Thus, we provide one mechanistic step toward understanding the Lyme disease emergence at northern latitudes of Europe.

Molecular characterization of colistin-resistant Klebsiella pneumoniae isolates and their conjugative mcr-carrying plasmids

BackgroundThis study aimed to characterize colistin-resistant K. pneumoniae (CoRKp) strains isolated from patients with urinary tract infections and bacteremia between 1999 and 2022 at a tertiary teaching hospital in Taiwan. MethodsA total of 1,966K. pneumoniae isolates were collected, among which 21 strains were identified as CoRKp. The antimicrobial susceptibility of these CoRKp strains to 19 antibiotics was assessed. The genome characteristics of 21 CoRKp strains were determined by Nanopore-Illumina hybrid whole genome sequencing. Additionally, conjugation assays were conducted to determine the transferability of plasmids carrying mcr genes to K. pneumoniae ATCC BAA-1706 and E. coli C600. The larvae infection model was used to analyze the differences in virulence between transconjugants and recipient strains. ResultsAmong the 21 CoRKp, 12 were multidrug-resistant, and four were extensively drug-resistant. The distribution of sequence types (STs) and K types among the CoRKp strains was quite diverse, and ST307 (5 strains) and K64 (3 strains) dominated in CoRKp. The insertion elements IS903B and ISVsa5, were found to inactivate mgrB of 1 and 2 CoRKp isolates, respectively. Moreover, 1, 4, 6, and 1 missense mutations of PhoQ, PmrA, PmrB, and MgrB, were identified in 21 CoRKp. Only two isolates SC-KP169 and SC-KP585 carried mcr-1 and mcr-8, respectively. The plasmid pSC-KP169-1 could be transferred inter- and intra-genus and contributed to the virulence of K. pneumoniae to larvae. In contrast, the plasmid pSC-KP585-1 could be transferred to E. coli but could not affect its virulence to larvae. ConclusionsWe identified 21 CoRKp from 1,966 isolates and found a conjugative plasmid carrying mcr-1 gene that contributed to the virulence of K. pneumoniae to larvae.

Endoparasites of Red Deer (Cervus elaphus L.) and Roe Deer (Capreolus capreolus L.) in Serbian Hunting Grounds.

In this study, parasitological examinations were conducted from 2019 to 2023. Fecal samples were collected from 289 wild ruminants (158 red deer and 131 roe deer) from hunting grounds in Vojvodina, which belong to the public company Vojvodinašume. Using qualitative and quantitative coprological diagnostic methods, the presence of protozoa (coccidia and Buxtonella sulcata), nematodes (gastrointestinal strongyles, Trichuris spp., Capillaria spp., Dictyocaulus spp., and Muellerius spp.), cestodes (Moniezia spp.), and trematodes (Fasciola hepatica, Fascioloides magna, Paramphistomum spp., and Dicrocoelium dendriticum) in the form of single and mixed infections were confirmed. Coproculture was used to identify the infective larvae of gastrointestinal strongyles. The total prevalence of endoparasitic infections in hunting ground 1 was 89.77% in red deer and 92.85% in roe deer, while in hunting ground 2, it was 72.97% in red deer and 85.96% in roe deer. Knowledge of the prevalence and assessment of the intensity of parasitic infections in wild ruminants is necessary for designing health protection programs in hunting grounds and planning control strategies, which are significant for this branch of hunting and public health.

The Effect of Parasitization by Trichodinid Ciliates on the Mortality of Cultured Pacific Fat Sleeper larvae (Dormitator latifrons).

Trichodinidae, a prevalent group of protozoan ectoparasites in aquaculture, cause rapid mortality in fish hatcheries. Despite their significance, knowledge about these parasites in farmed fish in South America, especially in native species that are currently being domesticated for aquaculture, remains limited. This study morphologically characterized the Trichodinid species that are infecting Pacific fat sleeper (Dormitator latifrons) larvae and evaluated their impact on larval rearing. Four pairs of broodstock were induced with GnRHA implants and placed in tanks containing 200 L of freshwater, with a water temperature of 28 ± 1.0 °C and a dissolved oxygen level of 4.00 ± 1.23 mg L-1, with partial water exchanges being performed daily. The larvae hatched 7 to 8 h after fertilization and were transferred to tanks containing water with the same quality parameters. Twelve hours post-hatching, the presence of Trichodina was observed. Every 24 h, 60 larvae per tank (n = 180 per treatment) were sedated, and larval wet mounts were prepared, air-dried at room temperature, and impregnated with silver nitrate. Infection parameters and daily mortality were calculated. Trichodina was observed to parasitize the pelvic fins, caudal fins, and heads of fish larvae, which showed lethargy and erratic swimming movements. The Trichodina species showed a daily increase in the infection parameters, and a 58% rate of larval mortality was observed at the fourth day post-hatching (dph) in the infected tanks. In captivity, D. latifrons larvae typically survive up to 7 days post-hatching (dph) before reaching their point of no return due to the lack of adequate diet and feeding regimes. However, our study indicates that Trichodina infestation accelerates mortality, causing infected larvae to die more quickly than uninfected ones.

Partial Protection of Goats against Haemonchus contortus Achieved with ADP-Ribosylation Factor 1 Encapsulated in PLGA Nanoparticles.

Haemonchus contortus (H. contortus), a nematode with global prevalence, poses a major threat to the gastrointestinal health of sheep and goats. In an effort to combat this parasite, a nanovaccine was created using a recombinant ADP-ribosylation factor 1 (ARF1) antigen encapsulated within poly lactic-co-glycolic acid (PLGA). This study aimed to assess the effectiveness of this nanovaccine in providing protection against H. contortus infection. Fifteen goats were randomly divided into three groups. The experimental group received two doses of the PLGA encapsulated rHcARF1 (rHcARF1-PLGA) nanovaccine on days 0 and 14. Fourteen days after the second immunization, both the experimental and positive control groups were challenged with 8000 infective larvae (L3) of H. contortus, while the negative control group remained unvaccinated and unchallenged. At the end of the experiment on the 63rd day, all animals were humanly euthanized. The results showed that the experimental group had significantly higher levels of sera IgG, IgA, and IgE antibodies, as well as increased concentrations of cytokines, such as IL-4, IL-9, IL-17, and TGF-β, compared to the negative control group after immunization. Following the L3 challenge, the experimental group exhibited a 47.5% reduction in mean eggs per gram of feces (EPG) and a 55.7% reduction in worm burden as compared to the positive control group. These findings indicate that the nanovaccine expressing rHcARF1 offers significant protective efficacy against H. contortus infection in goats. The results also suggest the need for more precise optimization of the antigen dose or a reassessment of the vaccination regimen. Additionally, the small sample size limits the statistical rigor and the broader applicability of the findings.

Rising temperatures may increase fungal epizootics in northern populations of the invasive spongy moth in North America

Insect pest species are generally expected to become more destructive with climate change because of factors such as weakened host tree defences during droughts and increased voltinism under rising temperatures; however, responses will vary by species due to a variety of factors, including altered interactions with their natural enemies. Entomopathogens are a substantial source of mortality in insects, but the likelihood of epizootics can depend strongly on climatic conditions. Previous research indicates that rates of infection of the spongy moth (Lymantria dispar) by its host-specific fungal pathogen, Entomophaga maimaiga, increase with environmental moisture and decrease as temperatures rise. High temperatures may have direct and indirect (due to the associated drying) effects on the fungus, but the interactive effects between temperature and moisture level on larval infection are unclear. Here, we test the hypothesis that warmer, drier conditions will decrease rates of infection of spongy moth larvae by E. maimaiga. We evaluated the effects of precipitation and temperature on larval mortality caused by E. maimaiga with a manipulative field experiment, conducted in one of the northernmost and coldest parts of the spongy moth’s non-native range in North America. We caged laboratory-reared spongy moth larvae in experimentally warmed open-air forest plots, exposing the larvae to soil inoculated with E. maimaiga resting spores during two consecutive trials. Caged larvae were exposed to three temperature treatments — ambient, 1.7 °C above ambient and 3.4 °C above ambient — and either supplemental precipitation (+173 mm per trial) or ambient precipitation. Opposite to our hypothesis, there was no significant effect of supplemental precipitation, nor an interaction between precipitation and temperature. There was, however, a significant positive effect of increasing temperature on the number of larvae infected. On average, in each respective trial, larval infection increased by 44% and 50% under the elevated temperature treatments compared to ambient temperature. Experimental warming may have increased infections because ambient temperatures at the field site were suboptimal for fungal germination. The results from this experiment suggest that, in colder portions of the spongy moth’s invasive range, increasing temperatures due to climate change may enhance the ability of E. maimaiga to help control populations of the spongy moth.

Transcriptomic response of Hermetia illucens L. (Diptera: Stratiomyidae) to wounding and Gram-negative bacterial infection

Abstract The larvae of the black soldier fly (BSFL), Hermetia illucens L. (Diptera: Stratiomyidae), are of economic interest due to their use as livestock feed component. Unraveling their response to a bacterial infection will allow us to gain a better insight into their biology. In the current study, we used RNA-Seq analysis to unravel the transcriptomic response of BSFL to wounding and infection by a Gram-negative bacterium, Pseudomonas protegens Pf-5. Five-day-old BSFL were subjected to three treatments, i.e. untreated, PBS-injection, and bacteria-injection (5000 CFU per larva) and samples were collected at three time points (2 h, 6 h, and 13 h) post- treatment. Wounding induced expression of genes encoding recognition molecules and signaling pathway genes such as PGRP-SA, and Relish, and antimicrobial peptides (AMPs) such cecropin, defensin, and attacin. At 2 h, wounding resulted in a significant upregulation of immunity-related genes whereas genes encoding for resilin and cuticle proteins were significantly downregulated. At 6 h, the expression of immunity-related genes reduced in response to wounding whereas their expression increased in infected larvae. At 13 h, the expression of immunity-related genes reduced drastically in response to wounding, while their expression increased significantly in infected larvae. Conversely, the expression of metabolism-related genes, such as trypsin and chymotrypsin, was significantly upregulated in wounded larvae at 13 h, while their expression was significantly reduced in infected larvae. Increased investments in immunity-related processes in infected BSF larvae correlated with the downregulation of genes associated with metabolic processes indicative of a trade-off. Various immunity-related genes, including those encoding cecropin, defensin, attacin, and PGRP-SA, were consistently induced only during pathogen infection, indicating their role in immunity against Gram-negative bacteria. In this study, we report multiple genes that are significantly upregulated post-bacterial infection in BSFL that may be utilized as biomarkers to monitor insect health in mass production facilities.

Revisiting immunity vs. exposure in schistosomiasis: A mathematical modeling study of delayed concomitant immunity.

The relative contributions of exposure vs. acquired immunity to the epidemiology of human schistosomiasis has been long debated. While there is considerable evidence that humans acquire partial immunity to infection, age- and sex-related contact patterns with water bodies contaminated with infectious cercarial schistosome larvae also contribute to typical epidemiological profiles of infection. Here, we develop a novel schistosome transmission model that incorporates both partially protective "delayed concomitant" acquired immunity-stimulated by dying worms-and host age- and sex-dependent patterns of exposure. We use a contemporary Bayesian approach to fit the model to historical individual data on exposure to infectious cercaria, eggs per gram of feces, and immunoglobulin E antibodies specific to Schistosoma mansoni Tegumental-Allergen-Like protein 1 collected from a highly endemic community in Uganda, estimating the relative contributions of exposure and acquired immunity. We find that model variants incorporating or omitting delayed concomitant immunity describe equally well the age- and sex-specific immunoepidemiological patterns observed before intervention and 18 months after treatment. Over longer time horizons, we find that acquired immunity creates subtle differences in immunoepidemiological profiles during routine mass drug administration that may confer resilience against elimination. We discuss our findings in the broader context of the immunoepidemiology of schistosomiasis.

Prevalence and distribution of trematodes hosted by Bithynia siamensis in Southern Thailand.

This study aimed to investigate the occurrence of larval trematode infections in bithyniid snails across five provinces in southern Thailand. A total of 1,413 Bithynia siamensis snails were collected between October 2021 and October 2022 using handpicking and scooping methods. Among these, 844 were identified as B. s. siamensis and 569 as B. s. goniomphalos. The snail samples were examined for parasitic infections in the laboratory using shedding and crushing methods. Among the 27 sampling sites, snails from 6 sites were infected with various trematode species (infection rate: 4.95%, 70/1,413). Evaluation of the morphological and internal organ characteristics of the cercariae allowed for the categorization of the trematodes into five species of trematodes (belonging to four families). The identified species included Echinochasmus pelecani (family: Echinostomatidae), Echinostoma revolutum (family: Echinostomatidae), Haematoloechus similis (family: Haematoloechidae), Loxogenoides bicolor (family Lecithodendriidae), and Stictodora tridactyla (family: Heterophyidae), and the infection rates for these species were 3.26% (46/1,413), 0.35% (5/1,413), 0.42% (6/1,413), 0.78% (11/1,413), and 0.14% (2/1,413), respectively. The cercariae from the identified trematode species were studied for DNA analysis. The phylogenetic lineage reveals relationships among the species, confirming the morphological distinctions.

Glucocorticoids Suppress NF-κB-Mediated Neutrophil Control of Aspergillus fumigatus Hyphal Growth.

Glucocorticoids are a major class of therapeutic anti-inflammatory and immunosuppressive drugs prescribed to patients with inflammatory diseases, to avoid transplant rejection, and as part of cancer chemotherapy. However, exposure to these drugs increases the risk of opportunistic infections such as with the fungus Aspergillus fumigatus, which causes mortality in >50% of infected patients. The mechanisms by which glucocorticoids increase susceptibility to A. fumigatus are poorly understood. In this article, we used a zebrafish larva Aspergillus infection model to identify innate immune mechanisms altered by glucocorticoid treatment. Infected larvae exposed to dexamethasone succumb to infection at a significantly higher rate than control larvae. However, both macrophages and neutrophils are still recruited to the site of infection, and dexamethasone treatment does not significantly affect fungal spore killing. Instead, the primary effect of dexamethasone manifests later in infection with treated larvae exhibiting increased invasive hyphal growth. In line with this, dexamethasone predominantly inhibits neutrophil function rather than macrophage function. Dexamethasone-induced mortality also depends on the glucocorticoid receptor. Dexamethasone partially suppresses NF-κB activation at the infection site by inducing the transcription of IκB via the glucocorticoid receptor. Independent CRISPR/Cas9 targeting of IKKγ to prevent NF-κB activation also increases invasive A. fumigatus growth and larval mortality. However, dexamethasone treatment of IKKγ crispant larvae further increases invasive hyphal growth and host mortality, suggesting that dexamethasone may suppress other pathways in addition to NF-κB to promote host susceptibility. Collectively, we find that dexamethasone acts through the glucocorticoid receptor to suppress NF-κB-mediated neutrophil control of A. fumigatus hyphae in zebrafish larvae.

Impacts of Pta-AckA pathway on CPS biosynthesis and type 3 fimbriae expression in Klebsiella pneumoniae

BackgroundKlebsiella pneumoniae is a Gram-negative bacterium that can cause infections, especially in individuals with diabetes. Recently, more hypervirulent strains have emerged, posing a threat even to healthy individuals. Understanding how K. pneumoniae regulates its virulence factors is crucial. Acetyl-phosphate (AcP) is essential for bacterial metabolism and can affect virulence factor expression. However, the role of the Pta-AckA pathway, which regulates AcP levels, in K. pneumoniae pathogenesis remains unclear. MethodsDeletion mutants lacking the pta and ackA, involved in AcP production and hydrolysis, were generated in K. pneumoniae CG43S3. Their effects on AcP levels, the patterns of global acetylated protein, capsular polysaccharide (CPS) amount, serum resistance, type 3 fimbriae expression, biofilm formation, and virulence in G. mellonella larva were assessed. ResultsDeletion of ackA in K. pneumoniae CG43S3 led to AcP accumulation, while pta deletion abolished AcP synthesis when grown in TB7+1 % glucose. This pathway influenced global protein acetylation, with pta deletion decreasing acetylation and ackA deletion increasing it. Additionally, pta deletion decreased the CPS amount, serum resistance, and type 3 fimbriae expression, while ackA deletion increased these factors. Furthermore, deleting pta and ackA attenuated the infected larva's virulence and death rate. ConclusionOur findings highlight the critical role of the Pta-AckA pathway in K. pneumoniae pathogenesis. This pathway regulates AcP levels, global protein acetylation, CPS production, serum resistance, and type 3 fimbriae expression, ultimately impacting virulence. The information provides insights into potential therapeutic targets for combating K. pneumoniae infection.

Assessment of housekeeping genes stability for gene transcription regulation analysis of Spodoptera littoralis (Lepidoptera: Noctuidae) under Spodoptera littoralis nucleopolyhedrovirus viral infection.

Normalization with respect to stable housekeeping genes is important to facilitate gene transcription regulation research and acquire more accurate quantitative polymerase chain reaction (qPCR) data. In the current study, five candidates housekeeping genes of the cotton leafworm, Spodoptera littoralis encoding for Actin (Actin), elongation factor 1-alpha (EF1α), ribosomal protein S3 (RPS3), ribosomal protein 49 (RP49), and Ubiquitin (Ubi), were evaluated as normalization housekeeping genes under Spodoptera littoralis nucleopolyhedrovirus (SpliNPV) viral infection. The qPCR results confirmed the expression of all five housekeeping genes in S. littoralis viral infected larvae. The expression profiles of the housekeeping genes showed that the EF1α, Actin, and RP49 had the minimum average Ct values of 18.41 ± 0.66, 18.84 ± 0.90 and 19.01 ± 0.87 in all infected samples, respectively. While RPS3 and Ubi showed the maximum average Ct of 21.61 ± 0.51 and 21.11 ± 0.82, respectively. According to the results of ΔCt and geNorm analysis, EF1α was ranked as the most stable housekeeping gene during infection time-course. While by using BestKeeper, geNorm and NormFinder, the Ubi, RP49, and RPS3 showed the most genes transcription stability. The obtained results were also validated using the Cytochrome c oxidase (COX) gene transcripts in response to SpliNPV infection. The results revealed that EF1α and Ubi were the most stable housekeeping genes to be used for normalizing S. littoralis gene transcription regulation under SpliNPV infection. These findings, provide a significant addition for gene transcription regulation studies of S. littoralis upon infection using SpliNPV as a bio-agent.

Chemical Control of Snail Vectors as an Integrated Part of a Strategy for the Elimination of Schistosomiasis-A Review of the State of Knowledge and Future Needs.

WHO promotes the implementation of a comprehensive strategy to control and eliminate schistosomiasis through preventive chemotherapy, snail control, clean water supply, improved sanitation, behaviour change interventions, and environmental management. The transmission of schistosomiasis involves infected definitive hosts (humans or animals) excreting eggs that hatch (miracidia), which infect freshwater snail vectors (also referred to as intermediate snail hosts) living in marshlands, ponds, lakes, rivers, or irrigation canals. Infective larvae (cercariae) develop within the snail, which, when released, may infect humans and/or animals in contact with the water. Snail control aims to interrupt the transmission cycle of the disease by removing the vector snails and, by so doing, indirectly improves the impact of the preventive chemotherapy by reducing reinfection. Snail control was, for many years, the only strategy for the prevention of schistosomiasis before preventive chemotherapy became the primary intervention. Snails can be controlled through various methods: environmental control, biological control, and chemical control. The chemical control of snails has proven to be the most effective method to interrupt the transmission of schistosomiasis. The current review aims to describe the vector snails of human schistosomiasis, present the chemicals and strategies for the control of snails, the challenges with the implementation, and the future needs. Snail control can play a key role in reducing schistosomiasis transmission and, thus, complements other interventions for disease control. There is a need to develop new molluscicide products or new formulations and methods of applications for existing molluscicides that would target snail vectors more specifically.

Anisakis extracellular vesicles elicit immunomodulatory and potentially tumorigenic outcomes on human intestinal organoids

BackgroundAnisakis spp. are zoonotic nematodes causing mild to severe acute and chronic gastrointestinal infections. Chronic anisakiasis can lead to erosive mucosal ulcers, granulomas and inflammation, potential tumorigenic triggers. How Anisakis exerts its pathogenic potential through extracellular vesicles (EVs) and whether third-stage infective larvae may favor a tumorigenic microenvironment remain unclear.MethodsHere, we investigated the parasite's tumorigenic and immunomodulatory capabilities using comparative transcriptomics, qRT-PCR and protein analysis with multiplex ELISA on human intestinal organoids exposed to Anisakis EVs. Moreover, EVs were characterized in terms of shape, size and concentration using classic TEM, SEM and NTA analyses and advanced interferometric NTA.ResultsAnisakis EVs showed classic shape features and a median average diameter of around 100 nm, according to NTA and iNTA. Moreover, a refractive index of 5–20% of non-water content suggested their effective biological cargo. After treatment of human intestinal organoids with Anisakis EVs, an overall parasitic strategy based on mitigation of the immune and inflammatory response was observed. Anisakis EVs impacted gene expression of main cytokines, cell cycle regulation and protein products. Seven key genes related to cell cycle regulation and apoptosis were differentially expressed in organoids exposed to EVs. In particular, the downregulation of EPHB2 and LEFTY1 and upregulation of NUPR1 genes known to be associated with colorectal cancer were observed, suggesting their involvement in tumorigenic microenvironment. A statistically significant reduction in specific mediators of inflammation and cell-cycle regulation from the polarized epithelium as IL-33R, CD40 and CEACAM1 from the apical chambers and IL-1B, GM-CSF, IL-15 and IL-23 from both chambers were observed.ConclusionsThe results here obtained unravel intestinal epithelium response to Anisakis EVs, impacting host’s anthelminthic strategies and revealing for the first time to our knowledge the host-parasite interactions in the niche environment of an emerging accidental zoonosis. Use of an innovative EV characterization approach may also be useful for study of other helminth EVs, since the knowledge in this field is very limited.Graphical

Thermosensory behaviors of the free-living life stages of Strongyloides species support parasitism in tropical environments.

Soil-transmitted parasitic nematodes infect over 1 billion people worldwide and are a common source of neglected disease. Strongyloides stercoralis is a potentially fatal skin-penetrating human parasite that is endemic to tropical and subtropical regions around the world. The complex life cycle of Strongyloides species is unique among human-parasitic nematodes in that it includes a single free-living generation featuring soil-dwelling, bacterivorous adults whose progeny all develop into infective larvae. The sensory behaviors that enable free-living Strongyloides adults to navigate and survive soil environments are unknown. S. stercoralis infective larvae display parasite-specific sensory-driven behaviors, including robust attraction to mammalian body heat. In contrast, the free-living model nematode Caenorhabditis elegans displays thermosensory behaviors that guide adult worms to stay within a physiologically permissive range of environmental temperatures. Do S. stercoralis and C. elegans free-living adults, which experience similar environmental stressors, display common thermal preferences? Here, we characterize the thermosensory behaviors of the free-living adults of S. stercoralis as well as those of the closely related rat parasite, Strongyloides ratti. We find that Strongyloides free-living adults are exclusively attracted to near-tropical temperatures, despite their inability to infect mammalian hosts. We further show that lifespan is shorter at higher temperatures for free-living Strongyloides adults, similar to the effect of temperature on C. elegans lifespan. However, we also find that the reproductive potential of the free-living life stage is enhanced at warmer temperatures, particularly for S. stercoralis. Together, our results reveal a novel role for thermotaxis to maximize the infectious capacity of obligate parasites and provide insight into the biological adaptations that may contribute to their endemicity in tropical climates.

211 United States Department of Agriculture National Animal Health Monitoring System Goat 2019 study: Regional prevalence of gastrointestinal nematodes and lungworms in the United States

Abstract One objective of the United States Department of Agriculture National Animal Health Monitoring System Goat 2019 study (September 2019-April 2020) was to determine the prevalence of gastrointestinal nematode (GIN) and lungworm infection in goats across the US. Feces were collected from 5,176 animals on 318 operations in 24 states and results were consolidated into 4 regions: Northwest (3 states, 34 operations), Northeast (9 states, 89 operations), Southeast (9 states, 132 operations) and Southwest (3 states, 63 operations). Feces from each operation were mixed and cultured to allow eggs to hatch and develop, The Baermann technique was used to recover infective third-stage larvae from cultures. GIN identification included Haemonchus, Trichostrongylus, Teladorsagia, Oesophagostomum and Nematodirus. Lungworm (Muellerius/Protostrongylus) first-stage larval presence was also noted. Frequencies were calculated and Haemonchus were found on 18.6 ± 3.5 % of the operations (Northwest, 27.0 ± 16.5 %; Northeast, 13.1 ± 4.7 %; Southeast, 15.9 ± 5.1 % and Southwest, 29.2 ± 9.4) %); Trichostrongylus on 97.2 ± 1.4 % of the operations (Northwest, 100 %; Northeast, 98±1.1 %; Southeast, 97.0 ± 2.2 % and Southwest, 95.7±4.2 %); Teladorsagia on 80.3 ± 4.0% of the operations (Northwest, 97.1 ± 2.4%; Northeast, 71.6 ± 9.1%; Southeast, 88.6 ± 4.6% and Southwest, 75.7 ± 7.3%); Oesophagostomum on 38.2 ± 4.3% of the operations (Northwest, 73.1 ± 9.2%; Northeast, 39.6 ± 7.1 %, Southeast, 46.4 ± 8.2% and Southwest, 15.5 ± 6.0%); Nematodirus on 7 ± 2.6 % of the operations (Northwest, 7 ± 5.1%; Northeast, 5.2 ± 4.7 %, Southeast, 0 % and Southwest, 20.8 ± 8.6 %); and lungworms on 35.4 ± 4.2 % of the operations (Northwest, 69.1 ± 9.8 %; Northeast, 52.2 ± 9.1 %, Southeast, 33.4±6.9% and Southwest, 8.3±3.3%). Results indicated that Trichostrongylus was the most prevalent followed by Teladorsagia, Oesophagostomum, lungworms, Haemonchus and Nematodirus. Prevalence of all GIN and lungworms were similar within regions except Nematodirus and lungworms, which were less in the southeast and southwest, respectively. Due to project seasonal timing and sampling logistics, fecal collection was conducted from fall through early spring which may have influenced prevalence, specifically the most pathogenic GIN, Haemonchus. This study provided valuable insight into USA regional prevalence of GIN and lungworms in goats.