Ascaris Infection Research Articles

Human Ascaris infection is associated with higher frequencies of IL-10 producing B cells.

Ascaris lumbricoides has dual effects on the immune system of infected hosts. The IgE response to this parasite has been thoroughly studied, but little is known about cellular responses induced by infection. This study aims to explore the interplay between A. lumbricoides infection and B cell responses, especially B regulatory cells. Participants from Santa Catalina, Bolívar, Colombia, a helminth-endemic town, were screened for soil-transmitted helminthiasis (STH) using stool examinations. Eighteen A. lumbricoides-infected and 11 non-infected subjects were selected. Blood samples were analyzed for Breg cells and related cytokines, and immunoglobulins specific to the A. lumbricoides excretory/secretory product, ABA-1. Infected subjects exhibited higher frequencies of Breg cells, especially those with a higher A. lumbricoides egg burden. Higher frequencies of different Breg subsets were observed in infected individuals, with CD25+CD71+CD73- B cells being notably increased in strongly infected individuals. Additionally, A. lumbricoides infection was associated with reduced levels of circulating ABA-1-specific IgG1 and IgE. IL-10+ B cell frequencies correlated inversely with ABA-1-specific IgE. A. lumbricoides infection has a significant impact on the immune response, particularly on Breg cell populations and antibody responses. Our findings suggest that A. lumbricoides infection mediates a dose-dependent immunosuppressive response characterized by an increase in Breg cells and concomitant suppression of ABA-1-specific humoral responses.

The concentration McMaster method for diagnosis of patent Ascaris and Trichuris infections in humans

In tropical and subtropical regions, soil-transmitted helminth (STH) infections such as Ascaris lumbricoides, Trichuris trichiura, and hookworms have a significant impact on public health. Globally, A. lumbricoides infects approximately 0.8 billion people, while T. trichiura infects around 500 million. This study involved a comparison of three diagnostic methods, Kato-Katz and two flotation methods (concentration McMaster, and simple McMaster), for the detection of Ascaris and Trichuris in human faeces. We conducted a comparison of the number of eggs in faeces (or faecal egg counts (FECs)) obtained with these methods using freshly collected samples that were positive for T. trichiura and spiked with a known quantity of Ascaris sp. eggs. Additionally, for the concentration McMaster method we assessed FECs after storing the samples at 5 °C for up to 21 days. The concentration McMaster method demonstrated superiority over the simple McMaster method in terms of higher detection levels for both helminths, while the Kato-Katz method yielded FEC values very close to the ’true’ spiking values. Importantly, the concentration McMaster method was considerably easier to read compared with the Kato-Katz method, and it allowed for sample storage at 5 °C for up to 14 days without affecting FEC results. Consequently, we conclude that the concentration McMaster method is an effective and user-friendly alternative for diagnosis of Ascaris and Trichuris infections in humans. Furthermore, it offers the advantage of sample storage before analysis, enhancing flexibility in the workflow.

Concurrent Ascaris infection modulates host immunity resulting in impaired control of Salmonella infection in pigs.

Ascaris is one of the most widespread helminth infections, leading to chronic morbidity in humans and considerable economic losses in pig farming. In addition, pigs are an important reservoir for the zoonotic salmonellosis, where pigs can serve as asymptomatic carriers. Here, we investigated the impact of an ongoing Ascaris infection on the immune response to Salmonella in pigs. We observed higher bacterial burdens in experimentally coinfected pigs compared to pigs infected with Salmonella alone. The impaired control of Salmonella in the coinfected pigs was associated with repressed interferon gamma responses in the small intestine and with the alternative activation of gut macrophages evident in elevated CD206 expression. Ascaris single and coinfection were associated with a rise of CD4-CD8α+FoxP3+ Treg in the lymph nodes draining the small intestine and liver. In addition, macrophages from coinfected pigs showed enhanced susceptibility to Salmonella infection in vitro and the Salmonella-induced monocytosis and tumor necrosis factor alpha production by myeloid cells was repressed in pigs coinfected with Ascaris. Hence, our data indicate that acute Ascaris infection modulates different immune effector functions with important consequences for the control of tissue-invasive coinfecting pathogens.IMPORTANCEIn experimentally infected pigs, we show that an ongoing infection with the parasitic worm Ascaris suum modulates host immunity, and coinfected pigs have higher Salmonella burdens compared to pigs infected with Salmonella alone. Both infections are widespread in pig production and the prevalence of Salmonella is high in endemic regions of human Ascariasis, indicating that this is a clinically meaningful coinfection. We observed the type 2/regulatory immune response to be induced during an Ascaris infection correlates with increased susceptibility of pigs to the concurrent bacterial infection.

Efficacy of Single-Dose Albendazole and Albendazole Plus Ivermectin for Soil-Transmitted Helminth Infection in Children in the Peruvian Amazon.

In countries where soil-transmitted helminth (STH) infections are endemic, deworming programs are recommended to reduce morbidity; however, increasing levels of resistance to benzimidazoles are of concern. In an observational study in Peru, we studied the clinical efficacy of 400 mg of albendazole 20 days after treatment among children aged 2-11 years. Of 426 participants who provided samples, 52.3% were infected with a STH, 144 (33.8%) were positive for Ascaris (41.8% light, 50.8% moderate, and 7.4% heavy infections), 147 (34.5%) were positive for Trichuris (75.2% light, 22.5% moderate, and 2.3% heavy infections), and 1.1% were positive for hookworm species (100% light infections). Additional stool samples were examined at 20, 90, and 130 days after the initial treatment. At 20 days post-administration of albendazole, the cure rate (CR) of Ascaris infection was 80.1% (95% CI: 73.5-86.7), and the egg reduction rate (ERR) was 70.8% (95% CI: 57.8-88.7); the CR for Trichuris infection was 27.1% (95% CI: 20.0-34.3), and the ERR was 29.8% (95% CI: -1.40 to 57.5). Among participants with persistent or recurrent infections with Trichuris, the combined therapy of albendazole (400 mg) and ivermectin at 600 µg/dose increased overall CR for Trichuris infection to 75.2% (95% CI: 67.3-83.2%) with an ERR of 84.2% (95% CI: 61.3-93.8%). Albendazole administration alone for the control of STH was associated with high rates of treatment failure, especially for Trichuris. Combined single doses of albendazole and ivermectin was observed to have improved efficacy.

Acute Ascaris infection impairs the effector functions of natural killer cells in single and Salmonella co-infected pigs

Natural killer (NK) cells play a key role in defense against Salmonella infections during the early phase of infection. Our previous work showed that the excretory/secretory products of Ascaris suum repressed NK activity in vitro. Here, we asked if NK cell functionality was influenced in domestic pigs during coinfection with Ascaris and Salmonella enterica serotype Typhimurium. Ascaris coinfection completely abolished the IL-12 and IL-18 driven elevation of IFN-γ production seen in CD16 + CD8α + perforin + NK cells of Salmonella single-infected pigs. Furthermore, Ascaris coinfection prohibited the Salmonella-driven rise in NK perforin levels and CD107a surface expression. In line with impaired effector functions, NK cells from Ascaris-single and coinfected pigs displayed elevated expression of the inhibitory KLRA1 and NKG2A receptors genes, contrasting with the higher expression of the activating NKp46 and NKp30 receptors in NK cells during Salmonella single infection. These differences were accompanied by the highly significant upregulation of T-bet protein expression in NK cells from Ascaris-single and Ascaris/Salmonella coinfected pigs. Together, our data strongly indicate a profound repression of NK functionality by an Ascaris infection which may hinder infected individuals from adequately responding to a concurrent bacterial infection.

Host gastric corpus microenvironment facilitates Ascaris suum larval hatching and infection in a murine model.

Ascariasis (roundworm) is the most common parasitic helminth infection globally and can lead to significant morbidity in children including chronic lung disease. Children become infected with Ascaris spp. via oral ingestion of eggs. It has long been assumed that Ascaris egg hatching and larval translocation across the gastrointestinal mucosa to initiate infection occurs in the small intestine. Here, we show that A. suum larvae hatched in the host stomach in a murine model. Larvae utilize acidic mammalian chitinase (AMCase; acid chitinase; Chia) from chief cells and acid pumped by parietal cells to emerge from eggs on the surface of gastric epithelium. Furthermore, antagonizing AMCase and gastric acid in the stomach decreases parasitic burden in the liver and lungs and attenuates lung disease. Given Ascaris eggs are chitin-coated, the gastric corpus would logically be the most likely organ for egg hatching, though this is the first study directly evincing the essential role of the host gastric corpus microenvironment. These findings point towards potential novel mechanisms for therapeutic targets to prevent ascariasis and identify a new biomedical significance of AMCase in mammals.

RARE CASE OF ASCARIASIS DETECTED BY COLONOSCOPY ON THE BACKGROUND OF ELEVATED LEVELS OF FECAL CALPROTECTIN

Introduction: The causative agents of ascariasis in humans are two species: Ascaris lumbricoides and Ascaris suum. For diagnosis, a fecal sample is most often examined. In some cases, the parasite can be identified when coming out with the intestinal passage, and very rarely up on colonoscopy. Aim to present a rare case of ascariasis where the diagnosis was made by colonoscopy on the background of elevated levels of fecal calprotectin (f-CP). Case presentation: A colonoscopy was performed on a 52-year-old female patient due to elevated f-CP. The patient had no complaints. The colonoscopy did not detect pathological changes of the intestinal mucosa, but documented larval stages of Ascaris spp. freely moving in the lumen of the large intestine. The patient was treated with albendazole. Subsequent parasitological examinations of fecal samples were negative. Discussion: In developed countries, the transmission of Ascaris lumbricoides is greatly reduced. On the background of a very limited transmission of Ascaris lumbricoides, many authors consider that most of the sporadic cases of ascariasis are due to Ascaris suum. In the case described by us, the f-CP levels normalized after the treatment, and for this reason, we cannot categorically reject the relationship between Ascaris infection and elevated f-CP levels. Conclusion: The presented clinical case is of interest due to the unusual way of diagnosi ascariasis. In the absence of clinical symptoms, and pathological changes of blood and biochemical parameters, except for elevated fecal calprotectin, inflammatory colon disease was suspected and was colonoscopy performed on this occasion.

The long-lasting Ascaris suum antigens in the lungs shapes the tissue adaptation modifying the pulmonary architecture and immune response after infection in mice

Ascariasis is the most prevalent helminth affecting approximately 819 million people worldwide. The acute phase of Ascariasis is characterized by larval migration of Ascaris spp., through the intestinal wall, carried to the liver and lungs of the host by the circulatory system. Most of the larvae subsequently transverse the lung parenchyma leading to tissue injury, reaching the airways and pharynx, where they can be expectorated and swallowed back to the gastrointestinal tract, where they develop into adult worms. However, some larvae are trapped in the lung parenchyma inciting an inflammatory response that causes persistent pulmonary tissue damage long after the resolution of infection, which returns to tissue homeostasis. However, the mechanism by which chronic lung disease develops and resolves remains unknown. Here, using immunohistochemistry, we demonstrate that small fragments and larval antigens of Ascaris suum are deposited and retained chronically in the lung parenchyma of mice following a single Ascaris infection. Our results reveal that the prolonged presence of Ascaris larval antigens in the lung parenchyma contributes to the persistent immune stimulation inducing histopathological changes observed chronically following infection, and clearly demonstrate that larval antigens are related to all phases of tissue adaptation after infection: lung injury, chronic inflammation, resolution, and tissue remodeling, in parallel to increased specific humoral immunity and the recovery of lung function in mice. Additional insight is needed into the mechanisms of Ascaris antigen to induce chronic immune responses and resolution in the host lungs following larval migration.

1H-NMR based-metabolomics reveals alterations in the metabolite profiles of chickens infected with ascarids and concurrent histomonosis infection

BackgroundGut infections of chickens caused by Ascaridia galli and Heterakis gallinarum are associated with impaired host performance, particularly in high-performing genotypes. Heterakis gallinarum is also a vector of Histomonas meleagridis that is often co-involved with ascarid infections. Here, we provide a first insight into the alteration of the chicken plasma and liver metabolome as a result of gastrointestinal nematode infections with concomitant histomonosis. 1H nuclear magnetic resonance (1H-NMR) based-metabolomics coupled with a bioinformatics analysis was applied to explore the variation in the metabolite profiles of the liver (N = 105) and plasma samples from chickens (N = 108) experimentally infected with A. galli and H. gallinarum (+H. meleagridis). This was compared with uninfected chickens at different weeks post-infection (wpi 2, 4, 6, 10, 14, 18) representing different developmental stages of the worms.ResultsA total of 31 and 54 metabolites were quantified in plasma and aqueous liver extracts, respectively. Statistical analysis showed no significant differences (P > 0.05) in any of the 54 identified liver metabolites between infected and uninfected hens. In contrast, 20 plasma metabolites including, amino acids, sugars, and organic acids showed significantly elevated concentrations in the infected hens (P < 0.05). Alterations of plasma metabolites occurred particularly in wpi 2, 6 and 10, covering the pre-patent period of worm infections. Plasma metabolites with the highest variation at these time points included glutamate, succinate, trimethylamine-N-oxide, myo-inositol, and acetate. Differential pathway analysis suggested that infection induced changes in (1) phenylalanine, tyrosine, and tryptophan metabolism, (2) alanine, aspartate and glutamate metabolism; and 3) arginine and proline metabolism (Pathway impact > 0.1 with FDR adjusted P-value < 0.05).ConclusionIn conclusion, 1H-NMR based-metabolomics revealed significant alterations in the plasma metabolome of high performing chickens infected with gut pathogens—A. galli and H. gallinarum. The alterations suggested upregulation of key metabolic pathways mainly during the patency of infections. This approach extends our understanding of host interactions with gastrointestinal nematodes at the metabolic level.

Malaria and soil transmitted helminths among patients attending some hospitals in Kano Metropolis, Kano State, Nigeria

Soil-transmitted helminths (STHs) and Plasmodium falciparum are known to the causative agents of neglected tropical diseases that continue to pose significant public health challenges in Nigeria and other endemic countries. This study was aimed at determining malaria and STHs coinfection in Kano metropolis, Kano state, Nigeria. The prevalence of malaria and STHs was determined using microscopy and formol ether concentration methods, respectively. Data on socio-demographic and risk factors were collected from three hundred (300) patients using structured questionnaires. The findings revealed a high prevalence of malaria (92.7%) and low STHs (19.0%). Malaria was more prevalent in males (52.3%) than females (40.3%), with the highest prevalence observed among individuals aged 6 – 12 years (25.7%). The difference in malaria prevalence based on age was not statistically significant (p&gt;0.05), contrary to gender which showed a significant difference in their association (p&lt;0.05). STHs infections were more common in participants aged between 6 - 12 years (11.3%), and the difference in prevalence among age groups was statistically significant (p&lt;0.05). Among the study sites, patients exhibited a higher prevalence of Ascaris infection (14.3%) compared to Hookworm infection (4.6%). Co-infection was recorded in 19.0% of the patients, while 73.7% had malaria infection alone. During this study, malaria and STHs were prevalent in Kano metropolis and there was a co-infection of Malaria and STHs among patients attending the selected hospitals in Kano.

Preschool child growth attainment and velocity during an agriculture intervention in rural Panama may be diminished by soil-transmitted helminths.

Agricultural interventions are often recommended to address undernutrition in subsistence farming communities. However, intensified agriculture exposure can increase soil transmitted helminth (STH) infections, which are linked with poor child growth. This study examined impacts of the VERASAN public health and agricultural intervention program on preschool child growth attainment (HAZ and WAZ) and relative growth velocity over 7 months [change in height-for-age (∆HAD) and weight-for-age (∆WAD) difference]. VERASAN was initiated in 15 subsistence farming communities in rural Panama experiencing chronic undernutrition. Activities targeted improved household food security, preschool child diets and growth by intensifying and diversifying household agriculture. Our objectives were to explore the relationship between VERASAN and preschool child growth attainment (HAZ and WAZ) and velocity (∆HAD and ∆WAD) during one agricultural cycle in 238 households. We compared those new to VERASAN with those involved for 1 or 5 years, and identified if agricultural practices, food security, diet diversity and treatment of pre-existing STH infection were associated with growth attainment or velocity. Prior participation in VERASAN did not directly influence WAZ, HAZ or ΔHAD but VERASAN-related benefits had an indirect influence. ΔHAD was positively associated with VERASAN-associated improvements in diet diversity and food security. HAZ and WAZ during land preparation were positively associated with diet diversity and HAZ with food security during harvest. HAZ was negatively associated with children visiting the agricultural plot, consuming leafy green vegetables and pre-existing hookworm infections. Both agricultural season and STH influenced ΔWAD. Children in VERASAN for 1 or 5 years experienced growth faltering between land preparation and growing season, but not those new to VERASAN. In contrast, between growing and harvest, ∆WAD declined in children new to VERASAN compared to children in VERASAN for longer. ΔWAD from land preparation to harvest was higher with pre-existing Ascaris infection whereas it was lower between growing season and harvest for pre-existing hookworm infection. In a context of preschool child growth faltering, malnutrition and STH infections, improved food security, agricultural production and diet diversity associated with VERASAN were associated with improved growth. In contrast, STH infections were negatively associated with some, but not all, growth outcomes.

Ascaris suum - A zoonosis in Bali, Indonesia.

Ascariasis is a common disease in humans and pigs. In previous reports, it has been known that ascariasis in humans is not only caused by Ascaris lumbricoides but also by Ascaris suum which is known to originate from pigs. This research aims to identify the species of Ascaris that infect traditional pig farmers in Bali, Indonesia. To achieve this, an observational study was conducted, in which 239 feces samples were collected from traditional pig farmers. The collected samples were then divided into two parts, with one part stored in a pot containing 10% formalin for microscopic testing, and the other part stored in 70% ethanol for molecular testing. In cases where Ascaris infection was observed during the microscopic examination, a further test was conducted using the polymerase chain reaction (PCR) method for molecular examination. The DNA derived from the PCR was then subjected to DNA sequencing. The result was compared to A. suum gene data previously published in the Gene Bank. The results of the microscopic examination showed that 22 farmers were infected with Ascaris. However, PCR testing and DNA sequencing indicated that three infections were caused by A. suum. Therefore, it can be concluded that A. suum is a zoonosis in Bali. Thus, efforts to control ascariasis must pay attention to the presence of pigs around it. Handling of ascariasis is not only carried out in humans but also in pigs and the environment.

A Rare Complication of Ascariasis: A Case of Acute Interstitial Nephritis.

Acute interstitial nephritis (AIN) due to helminths is a rare cause of acute kidney injury (AKI). Helminthiases often progresses insidiously, making diagnosis difficult. This was the case of a 72-year-old man, who presented with renal failure, itching and diarrhoea. Urinalysis revealed leukocyturia, microhaematuria and mild proteinuria. A full blood count revealed leucocytosis with eosinophilia. A stool parasitological examination revealed fertilised eggs of Ascaris lumbricoides. Tubulointerstitial nephropathy secondary to A. lumbricoides infection was suspected. A percutaneous renal biopsy was not performed since the patient refused the anti-platelet therapy discontinuation. Mebendazole, albendazole and prednisone therapy was administered. After worm eradiation and discharge, recovery from the parasitosis, absence of pruritus and eosinophilia, and progressive improvement of renal function were observed, strongly suggesting a causal relationship between Ascaris infection and AIN. Parasite infection should be considered in the differential diagnosis of unexplained renal failure because early diagnosis and treatment are necessary to avoid irreversible complications.

Pattern and repeatability of ascarid-specific antigen excretion through chicken faeces, and the diagnostic accuracy of coproantigen measurements as compared with McMaster egg counts and plasma and egg yolk antibody measurements in laying hens

BackgroundA coproantigen enzyme-linked immunosorbent assay (ELISA) has recently been proposed for detecting ascarid infections in chickens. The excretion pattern of ascarid antigens through chicken faeces and the consistency of measurements over the course of infections are currently unknown. This study evaluates the pattern and repeatability of worm antigen per gram of faeces (APG) and compares the diagnostic performance of the coproantigen ELISA with a plasma and egg yolk antibody ELISA and McMaster faecal egg counts (M-FEC) at different weeks post-infection (wpi).MethodsFaecal, blood and egg yolk samples were collected from laying hens that were orally infected with a mix of Ascaridia galli and Heterakis gallinarum eggs (N = 108) or kept as uninfected controls (N = 71). Measurements including (a) APG using a coproantigen ELISA, (b) eggs per gram of faeces (EPG) using the McMaster technique and (c) ascarid-specific IgY in plasma and in egg yolks using an ascarid-specific antibody ELISA) were performed between wpi 2 and 18.ResultsTime-dependent significant differences in APG between infected and non-infected laying hens were quantified. At wpi 2 (t(164) = 0.66, P = 1.00) and 4 (t(164) = −3.09, P = 0.094) no significant differences were observed between the groups, whereas infected hens had significantly higher levels of APG than controls by wpi 6 (t(164) = −6.74, P < 0.001). As indicated by a high overall repeatability estimate of 0.91 (CI = 0.89–0.93), APG could be measured consistently from the same individual. Compared to McMaster and antibody ELISA, coproantigen ELISA showed the highest overall diagnostic performance (area under curve, AUC = 0.93), although the differences were time-dependent. From wpi 6 to 18 coproantigen ELISA had an AUC > 0.95, while plasma IgY ELISA showed the highest diagnostic performance in wpi 2 (AUC = 0.95). M-FEC had the highest correlation with total worm burden, while APG had highest correlations with weights and lengths of A. galli.ConclusionAscarid antigen excretion through chicken faeces can be measured with high accuracy and repeatability using a coproantigen ELISA. The antigen excretion increases over time, and is associated with worm maturation, particularly with the size of A. galli. Our results suggest the necessity of complementary use of different diagnostic tools for a more accurate diagnosis of infections.Graphical

Intestinal Parasitic infection in Males of urban areas of district Bareilly in different age groups

Pathogenic protozoans, helminths, and nematodes are the primary reasons for diarrheal, vomiting, and stomach obstructions among babies and younger children. However, enteric helminths are now not viewed as serious sickness agents as the contamination is typically asymptomatic. The intestinal parasite's highest prevalence (34.78%) was recorded in the 5-15 years age group. The prevalence of Ascaris infection was highest (10%) in Urban areas of district Bareilly followed by Entamoeba (8.39%), pinworm (7.74%), and Giardia (3.87%) and hookworm (0.65%).

A copro-antigen ELISA for the detection of ascarid infections in chickens

A reliable method of diagnosing the most prevalent helminth infections in chickens is vital for developing effective control strategies. Ascaridia galli and Heterakisgallinarum are phylogenetically close nematode species that can elicit the development of cross-reactive antibodies in chickens. Therefore, an enzyme-linked immunoassay (ELISA) based on Ascaridia galli antigens in faeces of chickens to detect and quantify infections with both A. galli and H. gallinarum was developed. The ELISA utilised polyclonal antibodies that were obtained from rabbits immunised with soluble antigens isolated from A. galli. In two separate experiments, chickens were kept as uninfected controls or were orally infected with either 100 or 1000 of embryonated eggs of A. galli or H.gallinarum. Faecal samples were collected after 28–30 weeks post-infection. The ELISA was then used to quantify the concentration of soluble worm antigens in faecal samples, i.e., the amount of antigen per gram faeces, APG. The APG from infected chickens was significantly higher than non-infected groups in both experiments (P 0.001). Both 100 and 1000 infection dose groups were not significantly different (P = 0.999) in the experiment with H. gallinarum, whereas in the experiment with A. galli, APG was significantly higher in the 1000 infection group (P 0.001). A receiver operation characteristics (ROC) analysis that evaluates the qualitative performance of diagnostics tests was used to calculate the assay parameters within each mono-infection experiment. The result showed that the assay had a high diagnostics accuracy with an area-under-curve (AUC) of 0.99 in detecting infection in A. galli infected chickens and a moderate-high accuracy (AUC = 0.89) in birds infected with H. gallinarum. The diagnostic sensitivity and specificity of the assay at the optimal cut-off point equivalent to Youden index were 93% and 100% for detecting infections in A. galli experiment and 85% and 92% in H. gallinarum experiment, respectively. The correlation between faecal antigen concentration and all worm burden parameters was positive but generally low (r < 0.33), which provided less information about infection intensities. Nonetheless, these results indicate that a reliable and accurate qualitative diagnosis of the two most prevalent intestinal nematodes in chickens can be achieved using a non-invasive copro-antigen ELISA assay.

Identification of key interactions of benzimidazole resistance-associated amino acid mutations in Ascaris β-tubulins by molecular docking simulations

Ascaris species are soil-transmitted helminths that infect humans and livestock mainly in low and middle-income countries. Benzimidazole (BZ) class drugs have predominated for many years in the treatment of Ascaris infections, but persistent use of BZs has already led to widespread resistance in other nematodes, and treatment failure is emerging for Ascaris. Benzimidazoles act by binding to β-tubulin proteins and destabilising microtubules. Three mutations in the β-tubulin protein family are associated with BZ resistance. Seven shared β-tubulin isotypes were identified in Ascaris lumbricoides and A. suum genomes. Benzimidazoles were predicted to bind to all β-tubulin isotypes using in silico docking, demonstrating that the selectivity of BZs to interact with one or two β-tubulin isotypes is likely the result of isotype expression levels affecting the frequency of interaction. Ascaris β-tubulin isotype A clusters with helminth β-tubulins previously shown to interact with BZ. Molecular dynamics simulations using β-tubulin isotype A highlighted the key role of amino acid E198 in BZ-β-tubulin interactions. Simulations indicated that mutations at amino acids E198A and F200Y alter binding of BZ, whereas there was no obvious effect of the F167Y mutation. In conclusion, the key interactions vital for BZ binding with β-tubulins have been identified and show how mutations can lead to resistance in nematodes.

Efficient and Scalable Process to Produce Novel and Highly Bioactive Purified Cytosolic Crystals from Bacillus thuringiensis.

ABSTRACTBacillus thuringiensis (Bt) is a Gram-positive soil bacterium that is widely and safely applied in the environment as an insecticide for combatting insect pests that damage crops or are disease vectors. Dominant active ingredients made by Bt are insect-killing crystal (Cry) proteins released as crystalline inclusions upon bacterial sporulation. Some Bt Cry proteins, e.g., Cry5B (formally Cry5Ba1), target nematodes (roundworms) and show exceptional promise as anthelmintics (cures for parasitic nematode diseases). We have recently described inactivated bacteria with cytosolic crystal(s) (IBaCC) in which bioactive Bt Cry crystals (containing Cry5B) are fully contained within the cytosol of dead bacterial ghosts. Here, we demonstrate that these IBaCC-trapped Cry5B crystals can be liberated and purified away from cellular constituents, yielding purified cytosolic crystals (PCC). Cry5B PCC contains ~95% Cry5B protein out of the total protein content. Cry5B PCC is highly bioactive against parasitic nematode larvae and adults in vitro. Cry5B PCC is also highly active in vivo against experimental human hookworm and Ascaris infections in rodents. The process was scaled up to the 100-liter scale to produce PCC for a pilot study to treat two foals infected with the ascarid Parascaris spp. Single-dose Cry5B PCC brought the fecal egg counts of both foals to zero. These studies describe the process for the scalable production of purified Bt crystals and define a new and attractive pharmaceutical ingredient form of Bt Cry proteins.IMPORTANCE Bacillus thuringiensis crystal proteins are widely and safely used as insecticides. Recent studies have shown they also can cure gastrointestinal parasitic worm (nematode) infections when ingested. However, reproducible, scalable, and practical techniques for purifying these proteins have been lacking. Here, we address this severe limitation and present scalable and practical methods for large-scale purification of potently bioactive B. thuringiensis crystals and crystal proteins. The resultant product, called purified cytosolic crystals (PCC), is highly compatible with ingestible drug delivery and formulation. Furthermore, there are growing applications in agriculture and insect control where access to large quantities of purified crystal proteins is desirable and where these methods will find great utility.

Potentials and challenges in the isolation and detection of ascarid eggs in complex environmental matrices

Ascarid infections constitute a major concern for both human and animal health risk assessment. Although being effectively transmitted by soil, water and contaminated food, reliable detection of ascarid eggs in environmental media often remains challenging. However, contamination of the environment with ascarid ova has gained more attention as a decisive part of proper risk assessment in recent years. Due to various factors, such as sample matrices, dissociation detergents and flotation solutions, defined and standardised protocols for the isolation of eggs from complex environmental matrices are difficult to establish and therefore limited. Thus, this study reviews common techniques used for the recovery of ascarid eggs from environmental media with special emphasis on sampling strategies, purification procedures and microscopic as well as molecular detection of egg contamination. Despite various advancements, mainly in the field of molecular methods leading to more reliable and sensitive detection, it can be concluded that there is still a need for unified guidelines for sampling and recovery of ascarid eggs derived from complex environmental matrices.

TLR4 Gene Polymorphisms Interaction With Ascaris Infection in Severe RSV Bronchiolitis.

IntroductionThe identification of gene-environment interactions allows the recognition of groups with higher risk of morbidity. This study evaluated the interaction between the presence of TLR4 gene polymorphisms and Ascaris infection with severe bronchiolitis in a tropical Colombian region.MethodsWe included all infants younger than 24 months hospitalized due to bronchiolitis in Hospital centers in the county of Rionegro, Colombia. To identify interaction between severe bronchiolitis and presence of TLR4 polymorphisms and Ascaris infection, we used log-binomial regression.ResultsFour hundred and seventeen infants were hospitalized due to bronchiolitis, of which 115 (27%) had severe bronchiolitis. In infants with respiratory syncytial virus (RSV) acute infection and positive anti-Ascaris IgE, TLR4 Asp299Gly was associated to low risk of severe bronchiolitis (OR 0.09, CI 95% 0.01–0.48). Conversely, in infants RSV negative with negative anti-Ascaris IgE, TLR4 Asp299Gly was associated with an increased risk of severe bronchiolitis (OR 14.5, CI 95% 2.2–96).ConclusionIn our population there is an interaction between the presence of severe bronchiolitis, TLR4 Asp299Gly and Ile399Thr polymorphisms, anti-Ascaris IgE levels and RSV. This association should be evaluated in other populations to elucidate its role in the pathogenesis of severe bronchiolitis.