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Extensive backyard chickens are one of the most common production systems in low-income and middle-income countries (LMICs). In this production system, chickens are exposed to infectious forms of parasites as a result of the outdoor access and scavenging behaviour. Heterakis gallinarum is one of the most common nematode parasites present in the environment, and estimating its global and regional prevalence is essential for attributing the economic losses in extensive backyard chickens. The objective of this study is to predict the prevalence of heterakidosis in extensive backyard chickens at global and regional levels in LMICs using regression imputation methods. A binomial random effect model was developed using empirical data on heterakidosis prevalence and climatic factors as main predictors. Prevalence data were then imputed in all regions based on the regression model. Global and country prevalence were estimated based on regional predictions and their beta distributions. Minimum precipitation, minimum temperature and maximum temperature were selected as significant predictors. The population prevalence of heterakidosis was 0.24 (0.19-0.29). Countries with continental and dry climates had a higher mean prevalence, whereas countries with tropical climates had a lower mean prevalence of heterakidosis. As more empirical data on heterakidosis prevalence become available, this model and predictions should be redefined and updated to capture a more representative association and increase the accuracy of the predictions. The results of this study can be used to attribute the economic losses of extensive backyard chickens, taking into account a holistic approach as promoted by the GBADs programme and therefore, to identify which diseases are more costly to backyard farmers. Furthermore, results can be also served as a proxy for the risk of histomoniasis in extensive backyard chickens., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

During parasite infections, the liver may prioritise immune-related pathways over its metabolic functions. Intestinal infections caused by Ascaridia galli and Heterakis gallinarum impair feed intake, nutrient absorption, and weight gain. Histomonas meleagridis, vectored by H. gallinarum, can also damage liver tissues, potentially impairing liver functions. This study examined the hepatic gene expression in three strains of chickens: Ross-308 (R), Lohmann Brown Plus (LB), and Lohmann Dual (LD), 2 weeks after an experimental infection (n = 18) with both A. galli and H. gallinarum or kept as uninfected control (n = 12). Furthermore, H. gallinarum infection led to a co-infection with H. meleagridis. The mixed infections reduced feed intake and the average daily weight gain (P < 0.001). The infections also increased the plasma concentrations of alpha (1)-acid glycoprotein and the antibody titre against H. meleagridis (P = 0.049), with no strain differences (P > 0.05). For host molecular response, 1887 genes were differentially expressed in LD, while 275 and 25 genes were differentially expressed in R and LB, respectively. The up-regulated genes in R and LD were mostly related to inflammatory and adaptive immune responses, while down-regulated genes in LD were involved in metabolic pathways, including gluconeogenesis. Despite performance differences among the strains, worm burdens were similar, but hepatic molecular responses differed significantly. Moreover, there was an indication of a shift in hepatic functions towards immune-related pathways. We, therefore, conclude that the liver shifts its functions from metabolic to immune-related activities in chickens when challenged with mixed parasite species., (© 2024. The Author(s).)

Background: Heterakis gallinarum (H. gallinarum) is a common poultry parasite that can be found in the ceca of many gallinaceous bird species, causing minor pathology and reduced weight gain. Most infections go unnoticed in commercial flocks due to the dependence on fecal egg counts, which are prone to false-negative diagnoses. Furthermore, there is a lack of research on gastrointestinal nematodes that use molecular identification methods, which could be essential for rapid diagnosis and developing efficient control approaches. As a result, the study aimed to look at the cause of mortality in layer chickens induced by H. gallinarum in Egyptian poultry farms using morphological, ultrastructural, and molecular characterization. Histopathological, immunohistochemical, and cell-mediated immune responses from damaged cecal tissues were also examined., Results: Seventy bird samples from ten-layer flocks of different breeds (Native, white, and brown layers) suffering from diarrhea, decreased egg output, and emaciation were collected. Cecal samples were collected from affected and non-affected birds and were examined for parasitic diseases using light and a scanning electron microscope. The mitochondrial cytochrome oxidase 1 (COX1) gene was used to characterize H. gallinarum. Our results showed that the collected nematodal worms were identified as H. gallinarum (male and female), further confirmed by COX1 gene amplification and sequence alignment. Gene expression analysis of the inflammatory markers in infected tissues showed a significant up-regulation of IL-2, IFN-γ, TLR-4, and IL-1β and a significant down-regulation of the anti-inflammatory IL-10. The mRNA level of the apoptotic cas-3 revealed apoptotic activity among the H. gallinarum samples compared to the control group., Conclusions: Our results implemented the use of molecular methods for the diagnosis of Heterakis, and this is the first report showing the tissue immune response following infection in layers: upregulation of IL-1β, IFN-γ, Il-2, and TLR-4, while down-regulation of anti-inflammatory IL-10 in cecal tissue, Cas-3 apoptotic activity and Nuclear factor-κB (NF-κB)activity with immunophenotyping of T-cells in Heterakis infected tissue., (© 2024. The Author(s).)

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., Competing Interests: Conflict of interest The authors have read the journal’s policy and have the following competing interests: MH is an employee of TECODevelopment GmbH. This does not alter our adherence to Veterinary Parasitology policies on sharing data and materials. There is no patent associated with this research to declare. Other authors have no competing interest., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Free-range chickens are an integral part of poultry production in many developing countries. In the Mountain Kingdom of Lesotho, the majority of the population own free-range chickens, which serve a variety of purposes including being a source of meat, eggs and use for cultural rituals amongst others. However, there is lack of scientific studies on occurrence of parasitic infections on free-range chickens in Lesotho. The aim of this study was to document common gastrointestinal parasites infecting free-range chickens in four villages of Mafeteng District in Lesotho. A total number of 462 pooled faecal samples were collected from various households in HaKubutu (n = 114), HaMatjeka (n = 120), HaMpalipali (n = 120) and Thabang Villages (n = 108) which were subjected to microscopic examination using McMaster technique. The prevalence of gastrointestinal parasite infection was as follows: Eimeria tenella (12.8%), Ascaridia galli (10.4%) and Heterakis gallinarum (5%). The prevalence for H. gallinarum and Ascaridia galli were comparatively higher during the hot-wet season (7.1% and 2.8% respectively) than the cold-dry season (3.2% and 1.9% respectively) and varied significantly (P < 0.05). For E. tenella, the oocysts per gram were slightly higher in the cold-dry season than the hot-wet season. Polymerase chain reaction only amplified DNA from six (29%) adult A. galli worms of which two amplicons were successfully sequenced. The obtained cytochrome C oxidase subunit 1 partial gene sequences displayed 98-100% identity with South African A. galli isolates. This is the first scientific study on prevalence and molecular characterization of nematodes and coccidia species infecting free-range village chickens in Lesotho. The findings can be used to review management of gastrointestinal nematodes and protozoal parasites of free-range chickens in Lesotho., Competing Interests: Declaration of Competing Interest The authors have no conflicts of interest that might unduly influence their work., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Rural chicken production in Ghana is predominantly done under the extensive system that exposes birds to parasitic infections. We investigated the prevalence of Ascaridia spp. and Heterakis spp. and as a preliminary study characterized the genetic variance of the Ascaridia galli isolates from rural chicken in Kumbungu, Savelugu and Tolon Districts in the Northern Region, Ghana. A total of 86 chickens aged 6-10 weeks were dissected and GIT inspected for nematodes. Nematode were described based on morphological features to be A. galli and H. gallinarum. Additionally, the mitochondrial cox1 gene (475 bp) of Ascaridia isolates was amplified and sequenced. The overall prevalence of nematodes was 47.67%: A. galli 37.21% and H. gallinarum 20.93%. Prevalence values of A. galli in the Kumbungu, Savelugu and Tolon Districts were 25.00%, 36.00%, 56.00%, respectively, and that of H. gallinarum, respectively were 16.67%, 28.00% and 20.00%. A Chi-square test (x 2  = 6.0907, p < 0.048) showed an association of A. galli prevalence to the district of origin of birds. From 20 A. galli cox1 sequences analyzed, all sequences were identified as A. galli. Two haplotypes were recorded, namely, GHA1 and GHA2. Haplotype GHA1 was found to have wide distribution globally, whereas GHA2 appear to be novel in the present study. The data shows the importance of A. galli and H. gallinarum infection in rural chicken in northern Ghana and pave way for further epidemiological study of avian nematodes., (Copyright © 2022 Elsevier B.V. All rights reserved.)

The aim of this study was to describe the morphology and means of identification of helminths in native partridges (65) and introduced pheasants (32) in Poland and to determine the level of intestinal infection of these birds by helminths using parasitological and ecological indices. The birds were acquired during the hunting season in the years 2015-2017. Nematodes, Capillaria phasianina , cestodes, Railietina friedbergeri , and one trematode, Brachylaima sp. were recorded for the first time in partridges in Poland. Our findings indicate that parasites are more prevalent in pheasants (prevalence 70.4%) than in partridges (prevalence 50.0%). The component community and infracommunity of parasites of partridges are more diverse (Simpson's diversity index: 0.63 and mean Brillouin diversity index: 0.10 ± 0.17) and less dominated by a single parasite species ( Capillaria sp., Berger-Parker dominance index: 0.53) than the pheasant parasite community (Simpson's diversity index: 0.07, mean Brillouin diversity index: 0.005 ± 0.02, dominant species Heterakis gallinarum , Berger-Parker dominance index: 0.96). There were statistically significant differences between partridges and pheasants in the Brillouin diversity index and in the prevalence of Heterakis gallinarum (55.6% in pheasants vs. 19.0 in partridges). There were significant differences between wild and farmed partridges in the prevalence of infection by Capillaria sp. (4.3% vs. 37.5%) and H. gallinarum (39.1 vs. 6.2%). In conclusion, the pheasant was shown to be a reservoir, carrier, and shedder of nematodes, which may increase the risk of infection in partridges.

Heterakis gallinarum, H. beramporia, and H. indica are common nematodes in gallinaceous poultry in Asian countries, and the infections occasionally lead to declining health of the hosts. These three Heterakis spp. can be identified by the morphological characteristics of the male worms; however, the female worms and eggs cannot be identified because they have no reliable morphological characteristics for discrimination. In addition, H. gallinarum is a well-known vector of fetal protozoan Histomonas meleagridis, making the discrimination between these three Heterakis species important in basic and clinical veterinary parasitology. We analyzed nuclear ribosomal 18S-ITS1-5.8S-ITS2-28S DNA sequences of these three Heterakis species. The 18S, 5.8S, and 28S DNA sequences had very high homology between the species; however, the ITS1 and ITS2 sequence similarity was 68.5 %-93.2 %. H. gallinarum, H. beramporia, and H. indica were divided into separate clades in the ITS1 and ITS2-concatenated phylogenetic tree. Therefore, to develop a multiplex PCR method for discriminating between the three Heterakis species, we designed species-specific reverse primers within the ITS2 region as follows: H. gallinarum-specific HgI2-R, H. beramporia-specific HbI2-R5, and H. indica-specific HiI2-R. The multiplex PCR amplified 396-bp, 272-bp, and 482-bp fragments specific to H. gallinarum, H. beramporia, and H. indica DNA, respectively, and did not amplify the fragments using other chicken nematode DNAs such as Ascaridia galli, Oxyspirura mansoni, Dispharynx nasuta, and Cheilospirura hamulosa. These results suggest that the multiplex PCR would serve as a useful tool for identifying and diagnosing infections of H. gallinarum, H. beramporia, and H. indica in poultry., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The prevalence of Heterakis infection in reproductive geese from 56 flocks was investigated between February 2015 and November 2019 by using both anatomopathologic and coproscopic examinations. The nematodes were microscopically examined, and 37.5% (21/56) of the flocks were infected with Heterakis nematodes, mostly with Heterakis dispar (35.7%). In one particular flock, a Heterakis gallinarum infection was noted. In two flocks, Heterakis infection was identified only during coproscopic examination. The infection susceptibility of the geese depended on the bird's age-in the first reproductive season, 50% of the flocks were positive; in the second season, 42% were positive; and in the third season, 30% were positive. The number of nematodes in each necropsied goose varied from 8 to 216 individuals. During the 5 years of epidemiologic analysis, the infection occurrence decreased from 66.7% to 11%.

Heterakis gallinarum is a heavily prevalent poultry parasite that thrives in the ceca of various species of gallinaceous birds. It is a small roundworm, measuring between 4 and 15 mm long, in the family Heterakidae. Heterakis gallinarum has a direct life cycle not requiring an intermediate host to complete development, and it is generally believed that poultry raised at high density on litter are at greatest risk for accumulating large numbers of the nematode. This species typically only causes mild pathology that does not significantly affect bird performance. However, H. gallinarum is recognized as an economically important parasite by the poultry industry because its ovum serves as the vector for the protozoal parasite Histomonas meleagridis , the cause of histomonosis in poultry. Diagnosis of the nematode typically relies on fecal egg counts, which are prone to false negative diagnoses. Molecular tools are available for studying the nematode and diagnosing infected flocks. Treating and preventing H. gallinarum infection is made difficult due to the low efficacy of anthelmintics for eradicating H. gallinarum from infected birds and of disinfectants for destroying H. gallinarum ova on contaminated farms.

Nematodes are widespread and common in poultry. Disinfectants are used to reduce infection rates in poultry houses, but there is little documentation of their effectiveness. An in vitro assay was developed to test the efficacy of products to damage Heterakis gallinarum eggs, and nine disinfectants and chemicals commonly used in the poultry industry were tested. Embryonated eggs of H. gallinarum were pipetted into wells of plastic cell culture plates (250-300 eggs/well in water). Measured amounts of test articles were added to the suspensions for 2, 4, 6, or 24 hr. After exposure, eggs were washed with water and treated with trypan blue (1 ml of 0.4% solution, added to each well) at room temperature for 2 min. Eggshell integrity was determined microscopically by counting the number of eggs that were clear (intact) or that contained blue dye (compromised). As a test of embryo viability, five eggs per well from treatments containing compromised eggs were transferred to a Petri dish and hatched manually, using forceps to open the eggshell. Released larvae were then observed for signs of controlled movement. In a test of Clorox bleach (NaOCl), Green Klean, Decon7, Kem San, PLT, Virkon S, NaCl, dry limestone (CaCO 3 ), and diesel fuel, only NaOCl (bleach) and Green Klean damaged the eggshell, and only 20,625 ppm of NaOCl rendered the larvae nonviable.

Heterakis gallinarum is a widely distributed cecal nematode that parasitizes gallinaceous birds including chickens and turkeys. H. gallinarum infection poses a problem for the poultry industry as the nematode egg serves as a vector for the protozoan parasite, Histomonas meleagridis, the causative agent of histomonosis. The only means of detecting H. gallinarum in the environment is microscopic identification of the eggs in soil or feces; however, H. gallinarum eggs are often mistaken for those of Ascaridia galli. Three primer sets were designed from sequences cloned from the H. gallinarum genome to develop a diagnostic PCR. Each of these primer sets amplified a single product from H. gallinarum, but were unable to amplify DNA from H. meleagridis, Ascaridia galli, or Cestode sp. H. gallinarum DNA was amplified from Lumbricus sp. (earthworms) and Alphitobius diaperinus (darkling beetles), confirming that the earthworm acts as a paratenic host for H. gallinarum and suggesting that the darkling beetle may be a carrier for this nematode., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Introduction: Timely and valid diagnostic of parasitic diseases are prerequisites for profitable traditional poultry breeding, which enables adequate prophylaxis and effective therapy. This research is retrospective of the prevalence of endoparasites in 880 hens, 291 ducks, 171 geese and 302 turkeys in extensive poultry breeding in the Republic of Serbia., Materials and Methods: Qualitative parasitological examination was done by conventional gravitational flotation method and sedimentation method. In rare cases of animal death, an autopsy was performed., Results: In the period from 2012 to 2017, in 12 examined localities in the Republic of Serbia, single or mixed infections with endoparasites Eimeria spp., Ascaridia galli, Heterakis gallinarum, Capillaria contorta, Amidostomum anseris and Syngamus trachea were diagnosed. The highest prevalence of ascaridiosis was detected in hens (15.69-24.05%), trichostrongylidosis in ducks (20.53-30.19%), heterakiosis (20.89- 25.86%) and capillariosis (20.68-26.08%) in geese, and syngamosis (23.39%) and capillariosis (14.28- 24.17%) in turkeys. Endoparasites were the most prevalent in hens in south Serbia (63.21%), in ducks in central Serbia (54.71%) and in geese (46.55%) and turkeys (58.24%) in north Serbia. Among mixed infections predominant was polyparasitism of Heterakis spp. and Capillaria spp., Conclusions: An epizootiological and clinical diagnostic approach based on the results of parasitological screening is very important from the aspect of organic poultry farming.

Background: The nematophagous fungus Pochonia chlamydosporia can degrade ascarid (e.g. Ascaridia galli) eggs in agar and soil in vitro. However, it has not been investigated how this translates to reduced infection levels in naturally exposed chickens. We thus tested the infectivity of soil artificially contaminated with A. galli (and a few Heterakis gallinarum) eggs and treated with P. chlamydosporia. Sterilised and non-sterilised soils were used to examine any influence of natural soil biota., Methods: Unembryonated eggs were mixed with sterilised (S)/non-sterilised (N) soil, either treated with the fungus (F) or left as untreated controls (C) and incubated (22 °C, 35 days) to allow eggs to embryonate and fungus to grow. Egg number in soil was estimated on days 0 and 35 post-incubation. Hens were exposed to the soil (SC/SF/NC/NF) four times over 12 days by mixing soil into the feed. On day 42 post-first-exposure (p.f.e.), the hens were euthanized and parasites were recovered. Serum A. galli IgY level and ascarid eggs per gram of faeces (EPG) were examined on days -1 and 36 (IgY) or 40 p.f.e. (EPG)., Results: Egg recovery in SF soil was substantially lower than in SC soil, but recovery was not significantly different between NF and NC soils. SF hens had a mean worm count of 76 whereas the other groups had means of 355-453. Early mature/mature A. galli were recovered from SF hens whereas hens in the other groups harboured mainly immature A. galli. Heterakis gallinarum counts were low overall, especially in SF. The SF post-exposure IgY response was significantly lower while EPG was significantly higher compared to the other groups., Conclusions: Pochonia chlamydosporia was very effective in reducing ascarid egg numbers in sterilised soil and thus worm burdens in the exposed hens. However, reduced exposure of hens shifted A. galli populations toward a higher proportion of mature worms and resulted in a higher faecal egg excretion within the study period. This highlights a fundamental problem in ascarid control: if not all eggs in the farm environment are inactivated, the resulting low level infections may result in higher contamination levels with associated negative long-term consequences.

Heterakis gallinarum is one of the common parasitic nematodes found in the caecum of poultry. To investigate the genetic diversity and genetic structure of the H. gallinarum population in Sichuan, we amplified and sequenced the complete mitochondrial (mt) cytochrome c oxidase subunit II (cox2) gene of 59 H. gallinarum isolates from seven different geographical regions, then analyzed their genetic polymorphisms. All cox2 genes of the 59 H. gallinarum isolates were 696 bp in length, with an average A + T content of 67.1%. Fifty-nine sequences contained 34 variable sites, and were classified into 23 haplotypes (HS1-HS23). The values of haplotype diversity (Hd) and nucleotide diversity (π) were 0.688 and 0.00288, respectively. Based on values of F ST and Nm (F ST  = 0.01929, Nm = 12.71), there was a frequent gene flow but no significant genetic differentiation observed among the populations. The network map showed that the most prominent haplotype was HS1, and the other haplotypes (HS2-HS23) were centered on HS1 with a star-like topology, indicating that H. gallinarum had previously experienced a population expansion. To our knowledge, this is the first research on the population genetics of H. gallinarum based on mitochondrial cox2.

The nematodes Dispharynx spiralis (Superfamily-Spiruroidea, Family-Acuariidae) parasitising the proventriculus and Heterakis gallinarum (Superfamily-Subuluroidea, Family-Heterakidae) found in the caecum of two backyard poultry birds are described. The usual location of D. spiralis is glandular stomach or proventriculus, where their heads may be deeply buried in the proventricular wall. H. gallinarum occurs in the caecum and commonly called as caecal worm of poultry.

Population genetics information provides a foundation for understanding the transmission and epidemiology of parasite and, therefore, may be used to assist in the control of parasitosis. However, limited available sequence information in Heterakis gallinarum has greatly impeded the study in this area. In this study, we first investigated the genetic variability and genetic structure of H. gallinarum. The 1325 bp fragments of the mitochondrial COX1 gene were amplified in 56 isolates of H. gallinarum from seven different geographical regions in Sichuan province, China. The 56 sequences were classified into 22 haplotypes (H1-H22). The values of haplotype diversity (0.712) and nucleotide diversity (0.00158) in Sichuan population indicate a rapid expansion occurred from a relatively small, short-term effective population in the past. The haplotype network formed a distribution around H1 in a star-like topology, and the haplotypes did not cluster according to their geographical location. Similar conclusions could be made from MP phylogenetic tree. The Fst value (Fst<0.16965) and AMOVA analysis revealed that no significant genetic differentiation was observed among the seven different geographical populations. Neutrality tests (Tajima's D and Fu's Fs) and mismatch analysis indicated that H. gallinarum experienced a population expansion in the past. Our results indicated that H. gallinarum experienced a rapid population expansion in the past, and there was a low genetic diversity and an absence of population structure across the population.

Pheasants (Phasianus colchicus) and grey partridges (Perdix perdix) were maintained together on land known to be contaminated with eggs of the caecal nematode Heterakis gallinarum to examine the hypothesis that this shared parasite has a greater impact on grey partridges than on pheasants. Since an inverse relationship between worm intensity and partridge body condition was detected, while no such relationship with pheasant body condition was observed, we were unable to refute this hypothesis. Furthermore, that there was no relationship between worm intensity after the exposure period and partridge body mass prior to the infection trial implies that infection caused the decrease in partridge body condition, and not vice versa. Data consistent with previous observations that H. gallinarum fecundity and survival is greater in pheasants than in partridges suggest that the bulk source of nematode infection to wild grey partridges is reared pheasants, and not the partridges themselves. This, and the differential impact on host body condition, supports the hypothesis that the spread of parasites from increasing numbers of released pheasants has contributed to the decline in wild grey partridge populations in the UK within the past 50 years.