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Avian haemosporidian parasites are protozoans transmitted by vectors, belonging to the genera Plasmodium, Haemoproteus, and Leucocytozoon, capable of infecting various bird species. The prevalence of these parasites varies depending on factors such as the habitat and geographical location of the host and vector, as well as across different bird groups. Seabirds typically exhibit low infection rates due to the absence of vectors in marine environments. This study aimed to identify and characterize the molecular diversity of Haemosporida parasites in a population of Brown Boobies (Sula leucogaster) (n = 37) from the Santana Archipelago in Macaé, Rio de Janeiro state. We utilized PCR-based detection methods to confirm the presence of haemosporidian parasites and sequencing (ctyb gene) to delineate the lineages. The results uncovered a notable prevalence (48.6%) of haemosporidian parasites (Plasmodium and Haemoproteus), comprising 12 novel lineages and four previously documented lineages (establishing new host records). Our findings suggest that transmission and infection by these hemoparasites can occur both on and off the island. Overall, this study yields valuable insights into the genetic diversity of haemosporidian parasites in Brown Boobies, enriching our understanding of the ecology and epidemiology of these parasites in seabirds. [ABSTRACT FROM AUTHOR]

Simple Summary: African tropical regions have a remarkably high bird diversity, yet few studies have tried to unravel the presence of blood parasites in birds found in conservation areas. Knowing which blood parasites are present will help us to prepare for potential disease outbreaks. We test the hypothesis that conservation regions have a high diversity of parasites. Molecular methods were used to screen 1035 blood samples from 55 bird species for blood infections on sites inside and adjacent to the Kruger National Park in South Africa. Overall, 28.41% of birds were found infected with at least one type of blood parasites. Bird malaria of the type Haemoproteus and Plasmodium was found in 17.39% and 4.64% of the birds respectively. Leucocytozoon blood parasite was found in 9.24% of birds. One hundred distinct blood parasite types were detected, of which 56 were new types. Similar bird malaria (Haemoproteus and Plasmodium) infections were found in closely related birds, while Leucocytozoon was found in almost every bird type. Sites with a high bird diversity also had a high parasite diversity. These findings provide insight of how birds can gradually survive their blood parasite infections. Afrotropical regions have high bird diversity, yet few studies have attempted to unravel the prevalence of avian haemosporidia in conservation areas. The diversity and host specificity of parasites in biodiversity hotspots is crucial to understanding parasite distribution and potential disease emergence. We test the hypothesis that biodiverse regions are associated with highly diverse parasites. By targeting the cytochrome b (Cytb) gene, we molecularly screened 1035 blood samples from 55 bird species for avian haemosporidia infections to determine its prevalence and diversity on sites inside and adjacent to the Kruger National Park. Overall infection prevalence was 28.41%. Haemoproteus, Leucocytozoon, and Plasmodium presented prevalences of 17.39%, 9.24%, and 4.64%, respectively. One hundred distinct parasite lineages were detected, of which 56 were new lineages. Haemoproteus also presented the highest diversity compared to Leucocytozoon and Plasmodium with varying levels of specificity. Haemoproteus lineages were found to be specialists while Plasmodium and Leucocytozoon lineages were generalists. We also found a positive relationship between avian host diversity and parasite diversity, supporting an amplification effect. These findings provide insight data for host–parasite and co-evolutionary relationship models. [ABSTRACT FROM AUTHOR]

Background: Avian malaria is caused by diverse parasite species of the genus Plasmodium, and it affects various bird species. The occurrence of this disease in some wild bird species is sparsely documented due to the scarce availability of samples. Hence the pathogenicity in some hosts is not completely known. In addition, feral birds may act as reservoirs bridging the transmission cycle from wild migratory birds to domestic and zoo-kept bird species. Case presentation: An owner of pigeons adopted a feral pigeon (Columba livia forma domestica) and housed it together with his other pet-pigeons. The bird died unexpectedly a few weeks after a surgical procedure and necropsy revealed a severely anaemic carcass, with pale organs and hydropericardium. Histopathologic analysis revealed inflammatory infiltrates in the lung and liver, and monocytes and Kupffer cells contained haemozoin pigment indicative of phagocytosis of Plasmodium-infected erythrocytes. A high erythrocytic infection rate of 18% was evident in tissues and blood vessels in various organs. Furthermore, the thyroid had masses classified as thyroid carcinomas. Immunohistochemistry with anti- Plasmodium falciparum HSP70 antibody revealed positive signals in erythrocytes and intravascular leucocytes. Further microscopy analysis using a Hemacolor-stained impression smear revealed a high parasitaemia with an asynchronous infection showing all erythrocytic stages. Molecular diagnosis by PCR identified Plasmodium relictum, lineage GRW11 as the aetiological agent. The bird presented died most likely due to an acute infection as evidenced by the high blood parasitaemia, leading to major erythrocyte destruction. Further analyses of feral pigeons (n = 22) did not reveal any additional cases of Plasmodium infections. Conclusion: This study reports the first mortality associated with P. relictum lineage GRW11. The study supports previous studies, suggesting that Plasmodium infections are not frequent in pigeons. Host conditions like immunosuppression due to the tumour may have influenced the infection outcome in this fatal case. Use of anti-P. falciparum HSP70 antibody for detection of P. relictum antigens for immune assays in blood and tissue samples will be a useful tool for future studies. [ABSTRACT FROM AUTHOR]

Migrating animals are thought to be important spillover sources for novel pathogens. Haemosporidians (malaria‐related parasites) are one such group of pathogens that commonly spillover into novel host communities if competent vectors are present. In birds, shorebirds (sandpipers, plovers and allies) perform some of the longest avian migrations, yet they are traditionally perceived as relatively free from haemosporidians. Although low prevalence fits several theories, such as effective immune responses or low exposure to vectors, few studies have been carried out in freshwater inland sites, where the vectors of haemosporidians (e.g. mosquitoes) are abundant, with a mixture of actively migrating (staging) and breeding hosts. Here we report the prevalence of three haemosporidian parasites, Haemoproteus, Leucocytozoon and Plasmodium, screened in 214 shorebirds from 15 species sampled in a freshwater marshland, southern Belarus. Contrary to most previous studies, we found that haemosporidians were frequent, with an overall prevalence in the community of 16.36%, including the locally breeding shorebirds (23.13%, 134 individuals of 10 species). However, actively migrating shorebirds had much lower prevalence (0.05%, 55 individuals of five species). We suggest that blood parasite infections are more common in shorebirds than currently acknowledged. Yet, actively migrating species may be free from haemosporidians or carry suppressed infections, leading to lower prevalence or even apparent absence in some species. Taken together, we theorize that a combination of sampling biases has driven our understanding of haemosporidian prevalence in shorebirds and future studies should take the migratory status of individuals into account when reporting prevalence. Furthermore, we argue that birds undergoing active migration may be less likely sources of spillover events than previously assumed. [ABSTRACT FROM AUTHOR]

Abstract Background Avian malaria is caused by diverse parasite species of the genus Plasmodium, and it affects various bird species. The occurrence of this disease in some wild bird species is sparsely documented due to the scarce availability of samples. Hence the pathogenicity in some hosts is not completely known. In addition, feral birds may act as reservoirs bridging the transmission cycle from wild migratory birds to domestic and zoo-kept bird species. Case presentation An owner of pigeons adopted a feral pigeon (Columba livia forma domestica) and housed it together with his other pet-pigeons. The bird died unexpectedly a few weeks after a surgical procedure and necropsy revealed a severely anaemic carcass, with pale organs and hydropericardium. Histopathologic analysis revealed inflammatory infiltrates in the lung and liver, and monocytes and Kupffer cells contained haemozoin pigment indicative of phagocytosis of Plasmodium-infected erythrocytes. A high erythrocytic infection rate of 18% was evident in tissues and blood vessels in various organs. Furthermore, the thyroid had masses classified as thyroid carcinomas. Immunohistochemistry with anti- Plasmodium falciparum HSP70 antibody revealed positive signals in erythrocytes and intravascular leucocytes. Further microscopy analysis using a Hemacolor-stained impression smear revealed a high parasitaemia with an asynchronous infection showing all erythrocytic stages. Molecular diagnosis by PCR identified Plasmodium relictum, lineage GRW11 as the aetiological agent. The bird presented died most likely due to an acute infection as evidenced by the high blood parasitaemia, leading to major erythrocyte destruction. Further analyses of feral pigeons (n = 22) did not reveal any additional cases of Plasmodium infections. Conclusion This study reports the first mortality associated with P. relictum lineage GRW11. The study supports previous studies, suggesting that Plasmodium infections are not frequent in pigeons. Host conditions like immunosuppression due to the tumour may have influenced the infection outcome in this fatal case. Use of anti-P. falciparum HSP70 antibody for detection of P. relictum antigens for immune assays in blood and tissue samples will be a useful tool for future studies.

Haemosporidians are common blood parasites in wild bird populations, yet their effects on oxidative status remain understudied. Here, we measured the levels of thiobarbituric acid-reactive substances (TBARS) as an indicator of reactive oxygen species (ROS), total antioxidant capacity (TAC) as an indicator of non-enzymatic molecular antioxidants, and TBARS/TAC ratio as an indicator of oxidative status. We also used parasite genus-specific primers and PCR techniques to detect haemosporidians in 117 adults of Rufous-collared Sparrow (Zonotrichia capensis) from four locations in south–central Chile. Mixed-effect models were employed to compare oxidative indicators between infected and uninfected birds. Infected birds showed significantly higher TBARS levels, but no significant differences in TAC, leading to a higher TBARS/TAC ratio, especially in reproductive individuals. This suggests increased oxidative damage in infected birds, irrespective of sex or body condition. A positive relationship between TBARS and TAC was observed in both groups, but the antioxidant response was weaker in infected birds, indicating differential oxidative stress responses based on infection status. Body condition did not differ significantly between infected and uninfected individuals. These results demonstrate that haemosporidian infections impose oxidative costs on birds, potentially compounding the oxidative costs associated with reproduction.

Abstract Background The globally transmitted avian malaria parasite Plasmodium relictum (lineage SGS1) has been found to infect hundreds of different bird species with differences in infection outcomes ranging from more or less latent to potentially mortal. However, to date basic knowledge about the links between genetic differentiation and variation in infection outcome within this single malaria parasite species is lacking. Methods In this study, two different isolates of SGS1, obtained in the wild from two different host species, were used to investigate differences in their development in the blood and virulence in the experimentally infected canaries. Simultaneously, 258 kb of the parasite genome was screened for genetic differences using parasite mRNA and compared between experimental groups. Results The two isolates showed differences in development and caused mortality as well as effects on the blood parameters of their hosts. Although previous studies using single genes have shown very limited within lineage genetic diversity in the European population of SGS1, 226 SNPs were found across 322 genes, which separated the two experimental groups with a total of 23 SNPs that were fixed in either of the experimental groups. Moreover, genetic variation was found within each experimental group, hinting that each avian malaria infection harbours standing genetic variation that might be selected during each individual infection episode. Conclusion These results highlight extensive genetic variation within the SGS1 population that is transferred into individual infections, thus adding to the complexity of the infection dynamics seen in these host–parasite interactions. Simultaneously, the results open up the possibility of understanding how genetic variation within the parasite populations is linked to the commonly observed differences in infection outcomes, both in experimental settings and in the wild. Graphical Abstract

ABSTRACT Aim Location Methods Results Conclusions Changing biodiversity and environmental conditions may allow multi‐host pathogens to spread among host species and affect prevalence. There are several widely acknowledged theories about mechanisms that may influence variation in pathogen prevalence, including the controversially debated dilution effect and abundance‐occupancy relationship hypotheses. Here, we explore such abundance‐occupancy relationships for unique lineages of three vector‐borne avian blood parasite genera (the avian malaria parasite Plasmodium and the related haemosporidian parasites Parahaemoproteus and Leucocytozoon) across biogeographical regions.Nearctic‐Neotropical and Palearctic‐Afrotropical regions.We compiled a cross‐continental dataset of 17,116 bird individuals surveyed from 46 bird assemblages across the Nearctic‐Neotropical and Palearctic‐Afrotropical regions and explored relationships between local parasite lineage prevalence and host assemblage metrics in a Bayesian random regression framework.Most lineages from these three genera infected ≥ 5 host species and exhibited clear phylogenetic or functional host specificity. Lineage prevalence from all three genera increased with host range, but also with higher degrees of specialisation to phylogenetically or functionally related host species. Local avian community features were also found to be important drivers of prevalence. For example, bird species richness was positively correlated with lineage prevalence for Plasmodium and Leucocytozoon, whereas higher relative abundances of the main host species were associated with lower prevalence for Plasmodium and Parahaemoproteus but higher prevalence for Leucocytozoon.Our results broadly support several of the leading hypotheses about mechanisms that influence pathogen prevalence, including the niche breadth hypothesis in that higher avian host species diversity and broader host range amplify prevalence through increasing ecological opportunities and the trade‐off hypotheses in that specialisation among subsets of available host species may increase prevalence. Furthermore, the three studied avian haemosporidian genera exhibited different abundance‐occupancy relationships across the major global climate gradients and in relation to host availability, emphasising that these relationships do not strictly follow common rules for vector‐borne parasites with different life histories. [ABSTRACT FROM AUTHOR]

Simple Summary: This study explored how blood parasites, known as haemosporidians, impact the oxidative status of Rufous-collared Sparrows, a common bird species in Chile. Oxidative status measures the balance between harmful molecules called reactive oxygen species (ROS) and the bird's ability to neutralize them with antioxidants. The study found that birds infected with haemosporidians had higher levels of ROS, indicating more oxidative stress than uninfected birds. While the overall antioxidant levels did not differ between infected and uninfected birds, the infected birds showed a greater imbalance in ROS/TAC, suggesting they suffered more oxidative damage. This was especially true for birds in their reproductive phase. These findings suggest that haemosporidian infection, combined with the demands of reproduction, may impose significant health costs on these birds. Understanding this could help us learn more about how infections affect wildlife, and the strategies parasites use to survive in their hosts. Haemosporidians are common blood parasites in wild bird populations, yet their effects on oxidative status remain understudied. Here, we measured the levels of thiobarbituric acid-reactive substances (TBARS) as an indicator of reactive oxygen species (ROS), total antioxidant capacity (TAC) as an indicator of non-enzymatic molecular antioxidants, and TBARS/TAC ratio as an indicator of oxidative status. We also used parasite genus-specific primers and PCR techniques to detect haemosporidians in 117 adults of Rufous-collared Sparrow (Zonotrichia capensis) from four locations in south–central Chile. Mixed-effect models were employed to compare oxidative indicators between infected and uninfected birds. Infected birds showed significantly higher TBARS levels, but no significant differences in TAC, leading to a higher TBARS/TAC ratio, especially in reproductive individuals. This suggests increased oxidative damage in infected birds, irrespective of sex or body condition. A positive relationship between TBARS and TAC was observed in both groups, but the antioxidant response was weaker in infected birds, indicating differential oxidative stress responses based on infection status. Body condition did not differ significantly between infected and uninfected individuals. These results demonstrate that haemosporidian infections impose oxidative costs on birds, potentially compounding the oxidative costs associated with reproduction. [ABSTRACT FROM AUTHOR]

Simple Summary: Avian malaria, caused by Plasmodium, is considered to be a major vector-borne disease in raptors. Most infected raptors present no clinical manifestations, which are from subclinical to various morbidity. There are occasional reports of Plasmodium infected in raptors in Thailand. However, the information of parasite impact on infected raptors is limited. The objectives of this study are to investigate the prevalence in wild raptor populations by using active surveillance in several localities of Thailand and evaluate the impact of avian malaria in raptors' health. The authors found that Southern Thailand has higher prevalence of avian malaria than the other country's regions. This suggested that the raptors in Southern Thailand, which has humid environment, are more vulnerable to the malarial infection. Additionally, most tested raptors could tolerate the infection. The results from this study can be used for development of strategies for management of endangered raptor species, especially in rehabilitation institutes. Raptors (Accipitriformes, Falconiformes and Strigiformes) are important for ecological niches as bioindicators and an apex predator; however, their global populations have continuously decreased due to human activities, habitat loss and contagious diseases. Avian malaria that may cause the negative impact on raptors' health may also contribute to the declining of raptor populations. This study reported malaria's molecular prevalence and genetic diversity in wild-caught and rehabilitated raptors in the Kasetsart University Raptor Rehabilitation Unit. In total, 109 raptors from 18 provinces of Thailand were classified into two groups, which included 78 diurnal raptors (DIRs) in Accipitriformes and 31 nocturnal raptors (NORs) in Strigiformes. Each ethylenediaminetetraacetic (EDTA) blood sample (0.5–1 mL) was tested through haematological analyses and polymerase chain reaction (PCR)-based detection to assess parasites' health impacts. Amplicons of PCR positive samples were analysed for a nucleotide sequencing and phylogenetic relationships. The overall prevalence of avian malaria was low at 3.67% (4/109) (95% CI: 1.44–9.06%), with a prevalence of 3.86% (3/78) (95% CI: 1.32–10.70%) in DIRs and 3.23% (1/31) (95% CI: 1.32–10.70%) in NORs. Most of the infected samples were from southern Thailand. This suggested that the raptors in humid habitats are more vulnerable to the malarial infection, which was likely associated with vector and parasite abundance. Clinical appearances and haematological examinations demonstrated that raptors could tolerate the infection and only became asymptomatic and subclinically infected. This study is the first report of the infection of avian malaria in Cinerous Vulture, Himalayan Vulture and Barred Eagle Owl in Thailand, providing baseline information in preparedness for the disease diagnostic and further study of avian malaria in some endangered raptor species. [ABSTRACT FROM AUTHOR]

The role of plumage ornamentation as a signal of parasite infection is a key issue in the evolution of animal behaviour. Despite the undeniable role of host–parasite interactions in sexual selection, it is still unclear how parasites influence the variance in the expression of females displaying multiple ornaments. Here, by taking advantage of a long-term and individual-based, monitored population of pied flycatchers, Ficedula hypoleuca , in a Mediterranean area, we investigated the variation in the expression of wing patch (size) and forehead patch (occurrence and size) in relation to infections with haemosporidian parasites in breeding adult females. Haemosporidian infection was related to wing patch size, and this association varied in direction and magnitude depending on the origin of birds (locally born versus immigrant) and the type of breeding habitat (oakwood versus pine plantation). Specifically, differences in wing patch size were most evident among immigrant females breeding in the pine plantation, with noninfected females expressing larger wing patches than infected ones. Regarding the forehead patch, its occurrence was also modulated by haemosporidian infection through two-way interactions with bird origin (lower occurrence in locally born females) and breeding habitat type (lower occurrence among infected than uninfected females in the oakwood, whereas the opposite was true in the pine plantation). Only bird age predicted forehead patch size. Therefore, in addition to haemosporidian infection, the expression of these ornaments was related to extrinsic (breeding habitat) and intrinsic (age, origin) factors, which suggests the existence of trade-offs between physiological responses to infection and the expression of secondary sexual traits. Overall, our results suggest that plumage ornaments provide independent and reliable information on the haemosporidian infection status of female pied flycatchers. • Whether plumage ornaments are a sign of parasitic infections in females is unclear. • We assessed the links between two female ornaments and haemosporidian infection. • The expression of both ornaments was related to infection, origin, habitat and age. • However, the correlations between these factors differed depending on the ornament. • This suggests different signals for ornamentation – infection response trade-offs. [ABSTRACT FROM AUTHOR]

Background: The globally transmitted avian malaria parasite Plasmodium relictum (lineage SGS1) has been found to infect hundreds of different bird species with differences in infection outcomes ranging from more or less latent to potentially mortal. However, to date basic knowledge about the links between genetic differentiation and variation in infection outcome within this single malaria parasite species is lacking. Methods: In this study, two different isolates of SGS1, obtained in the wild from two different host species, were used to investigate differences in their development in the blood and virulence in the experimentally infected canaries. Simultaneously, 258 kb of the parasite genome was screened for genetic differences using parasite mRNA and compared between experimental groups. Results: The two isolates showed differences in development and caused mortality as well as effects on the blood parameters of their hosts. Although previous studies using single genes have shown very limited within lineage genetic diversity in the European population of SGS1, 226 SNPs were found across 322 genes, which separated the two experimental groups with a total of 23 SNPs that were fixed in either of the experimental groups. Moreover, genetic variation was found within each experimental group, hinting that each avian malaria infection harbours standing genetic variation that might be selected during each individual infection episode. Conclusion: These results highlight extensive genetic variation within the SGS1 population that is transferred into individual infections, thus adding to the complexity of the infection dynamics seen in these host–parasite interactions. Simultaneously, the results open up the possibility of understanding how genetic variation within the parasite populations is linked to the commonly observed differences in infection outcomes, both in experimental settings and in the wild. [ABSTRACT FROM AUTHOR]

Avian malaria is expanding upslope with warmer temperatures and driving multiple species of Hawaiian birds towards extinction. Methods to reduce malaria transmission are urgently needed to prevent further declines. Releasing Wolbachia-infected incompatible male mosquitoes could suppress mosquito populations and releasing Wolbachia-infected female mosquitoes (or both sexes) could reduce pathogen transmission if the Wolbachia strain reduced vector competence. We cleared Culex quinquefasciatus of their natural Wolbachia pipientis wPip infection and transinfected them with Wolbachia wAlbB isolated from Aedes albopictus. We show that wAlbB infection was transmitted transovarially, and demonstrate cytoplasmic incompatibility with wild-type mosquitoes infected with wPip from Oahu and Maui, Hawaii. We measured vector competence for avian malaria, Plasmodium relictum, lineage GRW4, of seven mosquito lines (two with wAlbB; three with natural wPip infection, and two cleared of Wolbachia infection) by allowing them to feed on canaries infected with recently collected field isolates of Hawaiian P. relictum. We tested 73 groups (Ntotal = 1176) of mosquitoes for P. relictum infection in abdomens and thoraxes 6–14 days after feeding on a range of parasitemias from 0.028% to 2.49%, as well as a smaller subset of salivary glands. We found no measurable effect of Wolbachia on any endpoint, but strong effects of parasitemia, days post feeding, and mosquito strain on both abdomen and thorax infection prevalence. These results suggest that releasing male wAlbB-infected C. quinquefasciatus mosquitoes could suppress wPip-infected mosquito populations, but would have little positive or negative impact on mosquito vector competence for P. relictum if wAlbB became established in local mosquito populations. More broadly, the lack of Wolbachia effects on vector competence we observed highlights the variable impacts of both native and transinfected Wolbachia infections in mosquitoes. Author summary: Avian malaria is expanding upslope in Hawaii with warmer temperatures and driving multiple species of birds towards extinction. Methods to reduce malaria transmission are urgently needed to prevent further declines. Releasing male mosquitoes infected with a type of bacteria (Wolbachia) could suppress mosquito populations by reducing offspring viability and releasing Wolbachia-infected female mosquitoes (or both sexes) could reduce pathogen transmission if the Wolbachia strain reduced vector competence. We cleared Culex quinquefasciatus of their natural Wolbachia pipientis wPip infection and transinfected them with Wolbachia wAlbB isolated from Aedes albopictus. We show that wAlbB infection was transmitted to offspring, and mating wAlbB males with wild-type mosquitoes produced mostly inviable offspring. We measured susceptibility to avian malaria infection of mosquitoes with native or transinfected Wolbachia (wPip or wAlbB). We found no measurable effect of Wolbachia on susceptibility. These results suggest that releasing male wAlbB-infected C. quinquefasciatus mosquitoes could suppress Hawaiian mosquito populations, but would have little positive or negative impact on mosquito vector competence for avian malaria. More broadly, the lack of Wolbachia effects on susceptibility to avian malaria we observed highlights the variable impacts of both native and transinfected Wolbachia infections in mosquitoes. [ABSTRACT FROM AUTHOR]

Parasites comprise a substantial portion of global biodiversity and play critical roles in shaping ecosystems by modulating trophic networks and affecting their hosts' abundance and distribution. The dynamics of host migration introduce new complexity to these relationships. From the host perspective, migratory behavior can either act as a defense mechanism or augment exposure to a broader spectrum of pathogens. Conversely, for parasites, host migration represents a mechanism for their dispersion and an opportunity to infect new host species. This study investigates the complex interplay between migration and parasite–host interactions, focusing on the interaction between hosts and avian malaria and malaria-like parasites in the Brazilian Atlantic Rain Forest. We captured 1466 birds representing 70 different species, uncovering 322 infections with Plasmodium/Haemoproteus parasites. We observed variations in migration timing and fluctuations in host abundance across months. By comparing the observed patterns of interaction of migratory and non-migratory birds to patterns of interaction expected at random, we show that migration affects the roles hosts take in the parasite–host network. Interestingly, despite the fact migratory species hosted more exclusive and distinct parasites, migrants did not occupy central network positions, which are mostly occupied by resident birds. Overall, we highlight the role of resident birds as a key species within parasite-host communities and the high specialization among avian haemosporidians and their hosts. [ABSTRACT FROM AUTHOR]

Host range expansions are an important factor for shaping the community of associated symbiotic organisms. Birds, as a highly mobile group of animals, are of particular interest to study with respect to the diversity and the distribution of the organisms using them as hosts during such large-scale movements. The Isabelline wheatear (Oenanthe isabellina) is a species with a main breeding area in Asia, which has expanded west, towards the Balkans, since the middle of the last century. We collected feather mites and blood samples for haemosporidian parasites screening from adult and juvenile Isabelline wheatears from the westernmost edge of the species breeding area in western Bulgaria. The feather mite species Alaudicola rosickyi (Černy, 1963), previously found on other wheatear species, was found for the first time on Isabelline wheatears. One species of avian malaria parasites—Plasmodium relictum (Grassi and Feletti, 1891) (lineage SGS1)—represented the first record of a malaria parasite in this host species in Europe. Increasing the sampling in the South European populations of the species will shed light on blood parasite species diversity and will reveal if other feather mite species have followed their host during its breeding range expansion. [ABSTRACT FROM AUTHOR]

Simple Summary: Aedes albopictus is an invasive mosquito species with a currently broad distribution range. In this article, we address the role of this mosquito species in the transmission of avian malaria parasites of the genus Plasmodium. To do that, we review the literature and compile existing information on Ae. albopictus–avian Plasmodium interactions to consider the potential impact of its expansion on avian malaria epidemiology. The findings suggest that while experimental infection of certain Plasmodium species is feasible, the occurrence of avian Plasmodium in wild Asian tiger mosquitoes is rare. Mosquitoes' preference for mammals as hosts and their relatively low susceptibility to infection may explain these results. The Asian tiger mosquito (Aedes albopictus) is an invasive mosquito species with a global distribution. This species has populations established in most continents, being considered one of the 100 most dangerous invasive species. Invasions of mosquitoes such as Ae. albopictus could facilitate local transmission of pathogens, impacting the epidemiology of some mosquito-borne diseases. Aedes albopictus is a vector of several pathogens affecting humans, including viruses such as dengue virus, Zika virus and Chikungunya virus, as well as parasites such as Dirofilaria. However, information about its competence for the transmission of parasites affecting wildlife, such as avian malaria parasites, is limited. In this literature review, we aim to explore the current knowledge about the relationships between Ae. albopictus and avian Plasmodium to understand the role of this mosquito species in avian malaria transmission. The prevalence of avian Plasmodium in field-collected Ae. albopictus is generally low, although studies have been conducted in a small proportion of the affected countries. In addition, the competence of Ae. albopictus for the transmission of avian malaria parasites has been only proved for certain Plasmodium morphospecies under laboratory conditions. Therefore, Ae. albopictus may play a minor role in avian Plasmodium transmission in the wild, likely due to its mammal-biased blood-feeding pattern and its reduced competence for the development of different avian Plasmodium. However, further studies considering other avian Plasmodium species and lineages circulating under natural conditions should be carried out to properly assess the vectorial role of Ae. albopictus for the Plasmodium species naturally circulating in its distribution range. [ABSTRACT FROM AUTHOR]

The major histocompatibility complex (MHC) is a genetic region in jawed vertebrates that contains key genes involved in the immune response. Associations between the MHC and avian malaria infections in wild birds have been observed and mainly explored in the Northern Hemisphere, while a general lack of information remains in the Southern Hemisphere. Here, we investigated the associations between the MHC genes and infections with Plasmodium and Haemoproteus blood parasites along a latitudinal gradient in South America. We sampled 93 rufous‐collared sparrows (Zonotrichia capensis) individuals from four countries, Colombia, Ecuador, Peru, and Chile, and estimated MHC‐I and MHC‐II allele diversity. We detected between 1–4 (MHC‐I) and 1–6 (MHC‐II) amino acidic alleles per individual, with signs of positive selection. We obtained generalized additive mixed models to explore the associations between MHC‐I and MHC‐II diversity and latitude. We also explored the relationship between infection status and latitude/biome. We found a non‐linear association between the MHC‐II amino acidic allele diversity and latitude. Individuals from north Chile presented a lower MHC genetic diversity than those from other locations. We also found an association between deserts and xeric shrublands and a lower prevalence of Haemoproteus parasites. Our results support a lower MHC genetic in arid or semi‐arid habitats in the region with the lower prevalence of Haemoproteus parasites. [ABSTRACT FROM AUTHOR]

Biodiversity varies worldwide and is influenced by multiple factors, such as environmental stability and past historical events (e.g. Panama Isthmus). At the same time, organisms with unique life histories (e.g. parasites) are subject to unique selective pressures that structure their diversity patterns. Parasites represent one of the most successful life strategies, impacting, directly and indirectly, ecosystems by cascading effects on host fitness and survival. Here, I focused on a highly diverse, prevalent and cosmopolitan group of parasites (avian haemosporidians) to investigate the main drivers (e.g. host and environmental features) of regional parasite diversity on a global scale. To do so, I compiled data from 4 global datasets on (i) avian haemosporidian (malaria and malaria-like) parasites, (ii) bird species diversity, (iii) avian functional traits and (iv) climate data. Then, using generalized least square models, I evaluated the effect of host and environmental features on haemosporidian diversity. I found that haemosporidian diversity mirrors host regional diversity and that higher host body mass increases haemosporidian diversity. On the other hand, climatic conditions had no effect on haemosporidian diversity in any model. When evaluating Leucocytozoon parasites separately, I found parasite diversity was boosted by a higher proportion of migratory hosts. In conclusion, I demonstrated that haemosporidian parasite diversity is intrinsically associated with their hosts’ diversity and body mass.

Endosymbiotic organelles, such as mitochondria and plastids, contain own remnant genomes (nucleoids), whose variable abundance in cells may be adaptive to the physiological necessities and functions of the cells. Unicellular apicomplexan parasites contain one mitochondrium and one apicoplast with variable genome copy numbers. We measured the abundance of mitochondrial, apicoplast and nuclear genome copies in a set of avian blood samples infected with haemosporidian blood parasites, belonging to the three main genera Plasmodium, Haemoproteus and Leucocytozoon. We designed general primers suitable for qPCR, amplifying fragments of the mitogenome, plastome and nuclear genomes of avian malaria and related haemosporidian parasites. We measured the amplification of these fragments in 153 samples of 23 avian host species and infected with 33 parasitic lineages. We estimate on average several hundred mitochondrial genome copies and several tens of apicoplast copies per haploid gametocyte cell with substantial variation among samples. Host species appeared to differ in their mitogenome abundance while parasitic lineages differed in plastome abundance per cell (per nuclear copy signal). We did not find consistent differences between parasite genera or higher avian taxa. Parasite lineages and host bird species did not differ consistently in infection intensity, estimated from parasite to host nuclear signals, which may indicate that samples were taken at different stages of infection. However, this and similar results remain to be cross-validated with in-situ imaging techniques. The novel molecular tools introduced here offer avenues for the characterization of nucleoid abundance of haemosporidian parasites over environmental conditions and parasitic developmental stages. Such measures will improve our understanding of parasite physiology, ecology, the coadaptation and coevolution with hosts and suggest possible augmentations to standard methods in the research field. [ABSTRACT FROM AUTHOR]

Illegal or poorly regulated wildlife trade may enhance parasite spread worldwide, leading to pathogen outbreaks and the emergence of diseases affecting native wildlife, domestic animals and humans. The order Psittaciformes has the largest proportion of endangered species among all birds worldwide and is one of the most trafficked taxa in the pet trade. However, despite the large number of parrot species commercialized worldwide, the influence of illegally traded wild birds on the introduction of exotic pathogens is still poorly investigated. Here we molecularly examined the prevalence and genetic diversity of haemosporidian parasites in illegally traded white‐winged parakeets (Brotogeris versicolurus), one of the most trafficked parrots in South America. We found that 18.5% of parakeets harboured Plasmodium relictum GRW04, a highly invasive malaria parasite provoking population decline and even extinctions in native avifauna when established outside its natural range. We also showed that malaria infected birds have lower body condition than uninfected parakeets, revealing the negative effects of malaria on their avian hosts. These outcomes highlight the risk of malaria spill over and disease outbreak in illegally traded wildlife. Our results also reveal epidemiological key concepts in disease transmission, such as the role of poorly studied parrot species as natural reservoir hosts of haemosporidians. These findings stress the importance of enforcing health control regulations and trade policies to fight wildlife trafficking effectively. [ABSTRACT FROM AUTHOR]

Infectious disease agents can influence each other's dynamics in shared host populations. We consider such influence for two mosquito-borne infections where one pathogen is endemic at the time that a second pathogen invades. We regard a setting where the vector has a bias towards biting host individuals infected with the endemic pathogen and where there is a cost to co-infected hosts. As a motivating case study, we regard Plasmodium spp., that cause avian malaria, as the endemic pathogen, and Usutu virus (USUV) as the invading pathogen. Hosts with malaria attract more mosquitoes compared to susceptible hosts, a phenomenon named vector bias. The possible trade-off between the vector-bias effect and the co-infection mortality is studied using a compartmental epidemic model. We focus first on the basic reproduction number R 0 for Usutu virus invading into a malaria-endemic population, and then explore the long-term dynamics of both pathogens once Usutu virus has become established. We find that the vector bias facilitates the introduction of malaria into a susceptible population, as well as the introduction of Usutu in a malaria-endemic population. In the long term, however, both a vector bias and co-infection mortality lead to a decrease in the number of individuals infected with either pathogen, suggesting that avian malaria is unlikely to be a promoter of Usutu invasion. This proposed approach is general and allows for new insights into other negative associations between endemic and invading vector-borne pathogens. [ABSTRACT FROM AUTHOR]

Avian haemosporidians of the genera Plasmodium and Haemoproteus are a group of widely distributed blood parasites that can negatively affect the fitness of their hosts. Colombia contains the greatest diversity of birds on the planet, but knowledge about the associations between haemosporidian and its avifauna is scarce and fragmented. We collected blood samples from 255 birds (203 residents and 52 neotropical migrants) belonging to 27 families and 108 species. The study was conducted in six localities in the inter-Andean valleys of the Cauca and Magdalena rivers. Parasites of the genera Plasmodium and Haemoproteus were identified in the samples by morphological and molecular analysis of a fragment of the mitochondrial gene cyt b. Among the samples, 9.3% (n = 24) were positive for Plasmodium or Haemoproteus. Co-infection with Plasmodium and Haemoproteus was found in Red-eyed Vireo. Seventeen haemosporidian lineages were identified, five of which were reported for the first time in resident birds (Common Ground Dove, Checker-throated Stipplethroat, Tropical Kingbird, Pale-breasted Thrush, and Ruddy-breasted Seedeater) and one in the Summer Tanager (neotropical migrant). The research results confirm the wide diversity of haemosporidian present in tropical lowlands and the possible role of neotropical migratory birds in dissemination on haemosporidian along their migratory routes. [ABSTRACT FROM AUTHOR]

Aim: Parasites play a fundamental role in shaping ecological communities and influencing trophic interactions. Understanding the factors that drive parasite impacts on community structure and stability (i.e. resilience to disturbances) is crucial for predicting disease dynamics and implementing effective conservation strategies. In this study, using avian malaria and malaria‐like parasites as a model system, we investigated the relationship between specificity, community stability and parasite vulnerability and their association with host diversity and climate. Location: Global. Time period: 2009–2023. Major taxa studied: Avian malaria and malaria‐like parasites. Methods: By compiling occurrence data from a global avian haemosporidian parasite database (MalAvi), we constructed a comprehensive dataset encompassing 60 communities. We utilized a phylogenetic model approach to predict missing host–parasite interactions, enhancing the accuracy of our analyses. Network analyses based on bipartite interactions provided measures of network specificity, stability, modularity, parasite competition and vulnerability to extinction. Results: We found that the high network specificity reduced community stability and decreased competition among parasites. Furthermore, we found that parasite vulnerability decreased with increasing community stability, highlighting the importance of community stability in host–parasite interactions for long‐term parasite persistence. When exploring the influence of local host diversity and climate conditions on host–parasite community stability, we demonstrated that increasing host biodiversity and precipitation reduces parasite competition. Conversely, higher temperature raises competition among parasites. Conclusion: These findings provide valuable insights into the mechanisms underlying parasite impacts on communities and the interplay between specificity, community stability and environmental factors. Further, we reveal the role of climate in shaping host–parasite interactions. By unravelling the complexities of parasite‐mediated interactions, our research substantially improves the current knowledge of the importance of specificity as a modulator of interactions in bipartite networks. [ABSTRACT FROM AUTHOR]

The story of the Hula Valley in the Galilee region of Palestine-Israel serves as the focus of this article, which draws on the concepts "more-than-One Health" and "settler ecologies" to highlight the harmful ecological implications of settler colonial projects in this region and elsewhere. Specifically, I tell the story of the Zionist drying of the Hula wetlands in the 1950s for the purpose of fighting off malaria and advancing agriculture in the region—and then of Israel's reflooding and rehabilitation of parts of the Hula in the 1990s in support of the massive annual bird migration. In winter 2021, more than eight thousand cranes succumbed to an avian influenza (H5N1) outbreak in the Hula Valley and over one million chickens in the area's coops had to be culled. Drawing on ethnographic fieldwork conducted mainly in summer 2022, this article discusses the historical and socioecological conditions that have arguably enabled and exacerbated the avian outbreak, advocating for a more-than-One Health approach that is founded on acknowledging the settler colonial legacies of this place. [ABSTRACT FROM AUTHOR]

Introduction: Bottleneck events are crucial for the strength of genetic drift, selection and speed of evolution. They are believed to play a particularly prominent role for parasitic infrapopulations, inhabiting single host individuals, which are often established by very few parasite individuals during transmission. In vector-borne pathogens, the bottlenecking effects can even be serialized through repeated filtering of parasitic stages at different tissues and organs of the vector. Using qPCR we aimed to quantify the number of potentially transmittable sporozoites of the hemosporidian blood parasite Haemoproteus columbae in the specialized vector louse flies Pseudolynchia canariensis which transmit these parasites between house pigeon hosts Columba livia. Results: Based on qPCR measurements of organ-derived DNA of individual louse flies, we estimate that the midgut of these vectors contains on average 20 parasites, the hindgut and other intestines ca. 50 parasites and the salivary glands ca. 5 parasite cells. Nearly one third of all vector individuals appeared to lack parasite DNA, despite having only infected hosts as blood meal sources. The magnitude of parasite numbers in midgut and salivary glands tended to correlate positively. Discussion: Our results indicate, potential severe bottlenecking of parasite populations during individual transmission events and a probable effect of individual vector immunity on this variable. However, this may be partly alleviated by the coloniality of house pigeons, the frequency of louse flies and their daily feeding events in most populations, leading to repeated transmission opportunities, decreased quasi-vertical transmission between parents and offspring and probable panmixia of Haemoproteus columbae lineages. Many of these mechanisms might not apply in other host-vector systems. We propose several additional molecular and microscopical tools to improve the accuracy of estimating parasite population sizes in vectors and call for more estimations in different vector species to better understand the co-evolution between malarialike blood parasites and their avian and insect hosts. [ABSTRACT FROM AUTHOR]

Patterns of pathogen prevalence are, at least partially, the result of coevolutionary host–pathogen interactions. Thus, exploring the distribution of host genetic variation in relation to infection by a pathogen within and across populations can provide important insights into mechanisms of host defence and adaptation. Here, we use a landscape genomics approach (Bayenv) in conjunction with genome‐wide data (ddRADseq) to test for associations between avian malaria (Plasmodium) prevalence and host genetic variation across 13 populations of the island endemic Berthelot's pipit (Anthus berthelotii). Considerable and consistent spatial heterogeneity in malaria prevalence was observed among populations over a period of 15 years. The prevalence of malaria infection was also strongly positively correlated with pox (Avipoxvirus) prevalence. Multiple host loci showed significant associations with malaria prevalence after controlling for genome‐wide neutral genetic structure. These sites were located near to or within genes linked to metabolism, stress response, transcriptional regulation, complement activity and the inflammatory response, many previously implicated in vertebrate responses to malarial infection. Our findings identify diverse genes – not just limited to the immune system – that may be involved in host protection against malaria and suggest that spatially variable pathogen pressure may be an important evolutionary driver of genetic divergence among wild animal populations, such as Berthelot's pipit. Furthermore, our data indicate that spatio‐temporal variation in multiple different pathogens (e.g. malaria and pox in this case) may have to be studied together to develop a more holistic understanding of host pathogen‐mediated evolution. [ABSTRACT FROM AUTHOR]

Simple Summary: Over thirty percent of bird species are undergoing population declines and are threatened with extinction in Portugal. Several reasons have been proposed to explain this decrease, such as the impact of human activities on natural environments and pathogens affecting the health of wildlife, domestic animals, and humans. Wildlife rescue and rehabilitation centres play an essential role in the conservation of endangered species. Despite wildlife rehabilitation centres providing valuable information on disease prevalence and transmission, the information on haemosporidian infection is still very scarce for birds admitted in these centres. In this study, we discovered new malaria parasites in birds admitted to wildlife rehabilitation centres in Portugal. We also revealed infection in bird species that were previously unknown to be infected with malaria parasites. Birds admitted to rehabilitation due to debilitating disease were more frequently infected with malaria. Furthermore, we demonstrate that the malaria infection extends the required period for medical treatment in these birds, which imposes additional economic costs for the rehabilitation and reduces the survival probabilities of the bird. These findings stress the importance of the study of malaria parasites in wildlife rehabilitation centres, also helping to design protocols and interventions to preserve endangered species. In the last decade, over 40% of bird species in Europe have experienced poor and bad conservation status, with more than 30% of bird species in mainland Portugal threatened with extinction. Along with anthropogenic factors, parasites and pathogens such as avian haemosporidians have been suggested to be responsible for these avian population declines. Wildlife rehabilitation centres play an essential role in species conservation and preservation. Moreover, animals admitted for rehabilitation can provide valuable information regarding transmission and pathogenicity of many diseases that affect wild birds that are rarely sampled in nature. However, reports of haemosporidians in captive birds are still limited. Here, we explored the prevalence and genetic diversity of avian haemosporidians in 89 birds from 29 species admitted to rehabilitation centres in Portugal, showing an overall infection prevalence of 30.3%. The prevalence of infection was higher in Strigiformes and in birds admitted to rehabilitation centres due to debilitating diseases. Remarkably, 30% of the infected bird species have not been found to harbour malaria parasites in preceding studies. We detected 15 different haemosporidian lineages infecting a third of bird species sampled. Notably, 2 out of these 15 detected haemosporidian lineages have not been obtained previously in other studies. Furthermore, we also identified nine new host–parasite interactions representing new host records for these haemosporidian parasites. Finally, our results revealed that birds infected with haemosporidians require longer rehabilitation treatments, which increase the economic costs for rehabilitation and may impair their survival prospects. These findings emphasise the importance of integrating haemosporidian infection considerations into rehabilitation protocols, highlighting the challenges posed by these infections in avian conservation and rehabilitation, including economic and logistical demands. [ABSTRACT FROM AUTHOR]

The ability of mosquitoes to transmit a pathogen is affected, among other factors, by their survival rate, which is partly modulated by their microbiota. Mosquito microbiota is acquired during the larval phase and modified during their development and adult feeding behavior, being highly dependent on environmental factors. Pharmaceutical residues including antibiotics are widespread pollutants potentially being present in mosquito breeding waters likely affecting their microbiota. Here, we used Culex pipiens mosquitoes to assess the impact of antibiotic exposure during the larval and adult stages on the survival rate of adult mosquitoes. Wild‐collected larvae were randomly assigned to two treatments: larvae maintained in water supplemented with antibiotics and control larvae. Emerged adults were subsequently assigned to each of two treatments, fed with sugar solution with antibiotics and fed only with sugar solution (controls). Larval exposure to antibiotics significantly increased the survival rate of adult females that received a control diet. In addition, the effect of adult exposure to antibiotics on the survival rate of both male and female mosquitoes depended on the number of days that larvae fed ad libitum in the laboratory before emergence. In particular, shorter larval ad libitum feeding periods reduced the survival rate of antibiotic‐treated adult mosquitoes compared with those that emerged after a longer larval feeding period. These differences were not found in control adult mosquitoes. Our results extend the current understanding of the impact of antibiotic exposure of mosquitoes on a key component of vectorial capacity, that is the vector survival rate. [ABSTRACT FROM AUTHOR]

Abstract Background The emergence of diseases of public health concern is enhanced by factors associated with global change, such as the introduction of invasive species. The Asian tiger mosquito (Aedes albopictus), considered a competent vector of different viruses and parasites, has been successfully introduced into Europe in recent decades. Molecular screening of parasites in mosquitoes (i.e. molecular xenomonitoring) is essential to understand the potential role of different native and invasive mosquito species in the local circulation of vector-borne parasites affecting both humans and wildlife. Methods The presence of avian Plasmodium parasites was molecularly tested in mosquitoes trapped in five localities with different environmental characteristics in southern Spain from May to November 2022. The species analyzed included the native Culex pipiens and Culiseta longiareolata and the invasive Ae. albopictus. Results Avian Plasmodium DNA was only found in Cx. pipiens with 31 positive out of 165 mosquito pools tested. None of the Ae. albopictus or Cs. longiareolata pools were positive for avian malaria parasites. Overall, eight Plasmodium lineages were identified, including a new lineage described here. No significant differences in parasite prevalence were found between localities or sampling sessions. Conclusions Unlike the invasive Ae. albopictus, Cx. pipiens plays a key role in the transmission of avian Plasmodium in southern Spain. However, due to the recent establishment of Ae. albopictus in the area, further research on the role of this species in the local transmission of vector-borne pathogens with different reservoirs is required. Graphical Abstract

Afrotropical regions have high bird diversity, yet few studies have attempted to unravel the prevalence of avian haemosporidia in conservation areas. The diversity and host specificity of parasites in biodiversity hotspots is crucial to understanding parasite distribution and potential disease emergence. We test the hypothesis that biodiverse regions are associated with highly diverse parasites. By targeting the cytochrome b (Cytb) gene, we molecularly screened 1035 blood samples from 55 bird species for avian haemosporidia infections to determine its prevalence and diversity on sites inside and adjacent to the Kruger National Park. Overall infection prevalence was 28.41%. Haemoproteus, Leucocytozoon, and Plasmodium presented prevalences of 17.39%, 9.24%, and 4.64%, respectively. One hundred distinct parasite lineages were detected, of which 56 were new lineages. Haemoproteus also presented the highest diversity compared to Leucocytozoon and Plasmodium with varying levels of specificity. Haemoproteus lineages were found to be specialists while Plasmodium and Leucocytozoon lineages were generalists. We also found a positive relationship between avian host diversity and parasite diversity, supporting an amplification effect. These findings provide insight data for host–parasite and co-evolutionary relationship models.

The correct identification of variables affecting parasite diversity and assemblage composition at different spatial scales is crucial for understanding how pathogen distribution responds to anthropogenic disturbance and climate change. Here, we used a database of avian haemosporidian parasites to test how the taxonomic and phylogenetic diversity and phylogenetic structure of the genera Plasmodium, Haemoproteus and Leucocytozoon from three zoogeographic regions are related to surrogate variables of Earth's energy input, habitat heterogeneity (climatic diversity, landscape heterogeneity, host richness and human disturbance) and ecological interactions (resource use), which was measured by a novel assemblage-level metric related to parasite niche overlap (degree of generalism). We found that different components of energy input explained variation in richness for each genus. We found that human disturbance influences the phylogenetic structure of Haemoproteus while the degree of generalism explained richness and phylogenetic structure of Plasmodium and Leucocytozoon genera. Furthermore, landscape attributes related to human disturbance (human footprint) can filter Haemoproteus assemblages by their phylogenetic relatedness. Finally, assembly processes related to resource use within parasite assemblages modify species richness and phylogenetic structure of Plasmodium and Leucocytozoon assemblages. Overall, our study highlighted the genus-specific patterns with the different components of Earth's energy budget, human disturbances and degree of generalism. [ABSTRACT FROM AUTHOR]

Background: Birds chronically infected with avian malaria parasites often show relapses of parasitaemia after latent stages marked by absence of parasites in the peripheral circulation. These relapses are assumed to result from the activation of dormant exo-erythrocytic stages produced during secondary (post-erythrocytic) merogony of avian Plasmodium spp. Yet, there is no morphological proof of persistent or dormant tissue stages in the avian host during latent infections. This study investigated persistence of Plasmodium relictum pSGS1 in birds with latent infections during winter, with the goal to detect presumed persisting tissue stages using a highly sensitive RNAscope® in situ hybridization technology. Methods: Fourteen domestic canaries were infected with P. relictum pSGS1 by blood-inoculation in spring, and blood films examined during the first 4 months post infection, and during winter and spring of the following year. After parasitaemia was no longer detectable, half of the birds were dissected, and tissue samples investigated for persisting tissue stages using RNAscope ISH and histology. The remaining birds were blood-checked and dissected after re-appearance of parasitaemia, and their tissues equally examined. Results: Systematic examination of tissues showed no exo-erythrocytic stages in birds exhibiting latent infections by blood-film microscopy, indicating absence of dormant tissue stages in P. relictum pSGS1-infected canaries. Instead, RNAscope ISH revealed rare P. relictum blood stages in capillaries of various tissues and organs, demonstrating persistence of the parasites in the microvasculature. Birds examined after re-appearance of parasitemia showed higher numbers of P. relictum blood stages in both capillaries and larger blood vessels, indicating replication during early spring and re-appearance in the peripheral circulation. Conclusions: The findings suggest that persistence of P. relictum pSGS1 during latent infection is mediated by continuous low-level erythrocytic merogony and possibly tissue sequestration of infected blood cells. Re-appearance of parasitaemia in spring seems to result from increased erythrocytic merogony, therefore representing recrudescence and not relapse in blood-inoculated canaries. Further, the study highlights strengths and limitations of the RNAscope ISH technology for the detection of rare parasite stages in tissues, providing directions for future research on persistence and tissue sequestration of avian malaria and related haemosporidian parasites. [ABSTRACT FROM AUTHOR]

Parasitic infections have the potential to impact the hosts' body condition, elevate physiological responses, and ultimately lead to increased mortality. Host-parasite interactions are tied to the ecological and life-history traits of the hosts. While montane birds are susceptible to avian blood parasites, few studies have simultaneously assessed how inter- and intra-specific traits of hosts influence their probability of parasite infection. In this study, we screened for avian blood parasites across 214 individuals from 51 species at two sites, including a lowland farmland at 700 m and a highland forest at 2,500 m, in the Gaoligong Mountains in southwestern China. Overall blood parasite prevalence was 53.74%, with divergent species-specific prevalence ranging from 6.25% to 66.67%. We also measured traits indicative of body condition and physiological responses of each sampled individual. Using Bayesian phylogenetic logistic models, we assessed whether parasite infection probability is associated with ecological and life history traits of host species. Larger bird species were more likely to be infected than smaller bird species, and omnivore species showed lower susceptibility than those with other diets such as insectivores and herbivores. In contrast, foraging strata, nest type, and participation in mixed-species flocks of host species did not affect infection probability. We then used a reduced sample of eight species with more than five individuals, to assess the associations between intra-specific infection probability and host body condition, represented by fat and muscle reserves, and acute stress responses measured through breath rate. While infected individuals were likely to have more fat reserves than non-infected individuals, we did not find any association between infection probability and muscle reserve and body mass, nor with breath rate. Our results revealed that at the species level, specific traits (body mass and diet) of host species predict infection probability and implied a potential link at the individual level between enhanced body condition and increased resilience to parasite infection. [ABSTRACT FROM AUTHOR]

Mosquitoes of the Culex pipiens complex are worldwide vectors of arbovirus, filarial nematodes, and avian malaria agents. In these hosts, the endosymbiotic bacteria Wolbachia induce cytoplasmic incompatibility (CI), i.e., reduced embryo viability in so-called incompatible crosses. Wolbachia infecting Culex pipiens (wPip) cause CI patterns of unparalleled complexity, associated with the amplification and diversification of cidA and cidB genes, with up to 6 different gene copies described in a single wPip genome. In wPip, CI is thought to function as a toxin-antidote (TA) system where compatibility relies on having the right antidotes (CidA) in the female to bind and neutralize the male's toxins (CidB). By repeating crosses between Culex isofemale lines over a 17 years period, we documented the emergence of a new compatibility type in real time and linked it to a change in cid genes genotype. We showed that loss of specific cidA gene copies in some wPip genomes results in a loss of compatibility. More precisely, we found that this lost antidote had an original sequence at its binding interface, corresponding to the original sequence at the toxin's binding interface. We showed that these original cid variants are recombinant, supporting a role for recombination rather than point mutations in rapid CI evolution. These results strongly support the TA model in natura, adding to all previous data acquired with transgenes expression. Long-term maintenance of Culex mosquito laboratory lines allowed this study to witness the emergence of novel cytoplasmic incompatibility patterns, showing that they arose from rapid evolution of Wolbachia genomes through gene loss and recombination. [ABSTRACT FROM AUTHOR]

[Display omitted] • A statistical relationship was created between quantitative PCR and microscopy to quantify Plasmodium relictum infection. • Infection detection probability differs between bird species due to differences in bird parasitemias. • Historical P. relictum (lineage GRW4) infection prevalence in Hawaii was likely much higher than previously thought. Plasmodium parasites infect thousands of species and provide an exceptional system for studying host-pathogen dynamics, especially for multi-host pathogens. However, understanding these interactions requires an accurate assay of infection. Assessing Plasmodium infections using microscopy on blood smears often misses infections with low parasitemias (the fractions of cells infected), and biases in malaria prevalence estimates will differ among hosts that differ in mean parasitemias. We examined Plasmodium relictum infection and parasitemia using both microscopy of blood smears and quantitative polymerase chain reaction (qPCR) on 299 samples from multiple bird species in Hawai'i and fit models to predict parasitemias from qPCR cycle threshold (Ct) values. We used these models to quantify the extent to which microscopy underestimated infection prevalence and to more accurately estimate infection patterns for each species for a large historical study done by microscopy. We found that most qPCR-positive wild-caught birds in Hawaii had low parasitemias (Ct scores ≥35), which were rarely detected by microscopy. The fraction of infections missed by microscopy differed substantially among eight species due to differences in species' parasitemia levels. Infection prevalence was likely 4–5-fold higher than previous microscopy estimates for three introduced species, including Zosterops japonicus , Hawaii's most abundant forest bird, which had low average parasitemias. In contrast, prevalence was likely only 1.5–2.3-fold higher than previous estimates for Himatione sanguinea and Chlorodrepanis virens , two native species with high average parasitemias. Our results indicate that relative patterns of infection among species differ substantially from those observed in previous microscopy studies, and that differences depend on variation in parasitemias among species. Although microscopy of blood smears is useful for estimating the frequency of different Plasmodium stages and host attributes, more sensitive quantitative methods, including qPCR, are needed to accurately estimate and compare infection prevalence among host species. [ABSTRACT FROM AUTHOR]

Stable isotope analysis provides valuable insights into the ecology of long-distance migratory birds during periods spent away from a specific study site. In a previous study, Swedish great reed warblers (Acrocephalus arundinaceus) infected with haemosporidian parasites differed in feather isotope ratios compared to non-infected birds, suggesting that infected and non-infected birds spent the non-breeding season in different locations or habitats. Here, we use a novel dataset comprising geolocator data, isotopes, and haemosporidian infection status of 92 individuals from four Eurasian populations to investigate whether parasite transmission varies with geography or habitats. We found that the probability of harbouring Plasmodium and Leucocytozoon parasites was higher in birds moulting in the eastern region of the non-breeding grounds. However, no geographic pattern occurred for Haemoproteus infections or overall infection status. In contrast to the previous study, we did not find any relationship between feather isotope ratios and overall haemosporidian infection for the entire current dataset. Plasmodium-infected birds had lower feather δ15N values indicating that they occupied more mesic habitats. Leucocytozoon-infected birds had higher feather δ34S values suggesting more coastal sites or wetlands with anoxic sulphate reduction. As the composition and prevalence of haemosporidian parasites differed between the old and the current dataset, we suggest that the differences might be a consequence of temporal dynamics of haemosporidian parasites. Our results emphasize the importance of replicating studies conducted on a single population over a restricted time period, as the patterns can become more complex for data from wider geographical areas and different time periods. [ABSTRACT FROM AUTHOR]

In a 2-yr study on prevalence of Haemosporidia in an avian community in Ithaca, New York, USA, we tested the hypothesis that apparent seasonal variation in prevalence is influenced by the detection protocol. We confirmed a higher detection of Haemosporidia using a molecular diagnosis technique (PCR) than by microscopy; this further increased when the PCR test was triplicated. Microscopic examination and PCR techniques have different specificity and sensitivity and therefore different probabilities of detecting hemoparasites. Birds with chronic infections or sampled during winter often have very low parasitemia, and such infections may be missed by microscopy but detected by PCR. Haemosporidian prevalence was higher during the breeding season than during the nonbreeding season regardless of the method used. Detection of Leucocytozoon spp. infection from blood smears using microscopy was challenging. [ABSTRACT FROM AUTHOR]

Avian malaria infection has been known to affect host microbiota, but the impact of Plasmodium infection on the colonization resistance in bird gut microbiota remains unexplored. This study investigated the dynamics of Plasmodium relictum infection in canaries, aiming to explore the hypothesis that microbiota modulation by P. relictum would reduce colonization resistance. Canaries were infected with P. relictum, while a control group was maintained. The results revealed the presence of P. relictum in the blood of all infected canaries. Analysis of the host microbiota showed no significant differences in alpha diversity metrics between infected and control groups. However, significant differences in beta diversity indicated alterations in the microbial taxa composition of infected birds. Differential abundance analysis identified specific taxa with varying prevalence between infected and control groups at different time points. Network analysis demonstrated a decrease in correlations and revealed that P. relictum infection compromised the bird microbiota's ability to resist the removal of taxa but did not affect network robustness with the addition of new nodes. These findings suggest that P. relictum infection reduces gut microbiota stability and has an impact on colonization resistance. Understanding these interactions is crucial for developing strategies to enhance colonization resistance and maintain host health in the face of parasitic infections. [ABSTRACT FROM AUTHOR]

IntroductionBottleneck events are crucial for the strength of genetic drift, selection and speed of evolution. They are believed to play a particularly prominent role for parasitic infrapopulations, inhabiting single host individuals, which are often established by very few parasite individuals during transmission. In vector-borne pathogens, the bottlenecking effects can even be serialized through repeated filtering of parasitic stages at different tissues and organs of the vector. Using qPCR we aimed to quantify the number of potentially transmittable sporozoites of the hemosporidian blood parasite Haemoproteus columbae in the specialized vector louse flies Pseudolynchia canariensis which transmit these parasites between house pigeon hosts Columba livia.ResultsBased on qPCR measurements of organ-derived DNA of individual louse flies, we estimate that the midgut of these vectors contains on average 20 parasites, the hindgut and other intestines ca. 50 parasites and the salivary glands ca. 5 parasite cells. Nearly one third of all vector individuals appeared to lack parasite DNA, despite having only infected hosts as blood meal sources. The magnitude of parasite numbers in midgut and salivary glands tended to correlate positively.DiscussionOur results indicate, potential severe bottlenecking of parasite populations during individual transmission events and a probable effect of individual vector immunity on this variable. However, this may be partly alleviated by the coloniality of house pigeons, the frequency of louse flies and their daily feeding events in most populations, leading to repeated transmission opportunities, decreased quasi-vertical transmission between parents and offspring and probable panmixia of Haemoproteus columbae lineages. Many of these mechanisms might not apply in other host-vector systems. We propose several additional molecular and microscopical tools to improve the accuracy of estimating parasite population sizes in vectors and call for more estimations in different vector species to better understand the co-evolution between malaria-like blood parasites and their avian and insect hosts.

Avian haemosporidians are protozoan parasites transmitted by insect vectors that infect birds worldwide, negatively impacting avian fitness and survival. However, the majority of haemosporidian diversity remains undescribed. Quantifying this diversity is critical to determining parasite–host relationships and host-switching potentials of parasite lineages as climate change induces both host and vector range shifts. In this study, we conducted a community survey of avian haemosporidians found in breeding birds on the Davis Mountains sky islands in west Texas, USA. We determined parasite abundance and host associations and compared our results to data from nearby regions. A total of 265 birds were screened and infections were detected in 108 birds (40.8%). Most positive infections were identified as Haemoproteus (36.2%), followed by Plasmodium (6.8%) and Leucocytozoon (0.8%). A total of 71 haemosporidian lineages were detected of which 39 were previously undescribed. We found that regional similarity influenced shared lineages, as a higher number of lineages were shared with avian communities in the sky islands of New Mexico compared to south Texas, the Texas Gulf Coast and central Mexico. We found that migratory status of avian host did not influence parasite prevalence, but that host phylogeny is likely an important driver.

The relationships between host phylogenetics, functional traits and parasites in wildlife remain poorly understood in the Neotropics, especially in habitats with marked seasonal variation. Here, we examined the effect of seasonality and host functional traits on the prevalence of avian haemosporidians (Plasmodium and Haemoproteus) in the Brazilian Caatinga, a seasonally dry tropical forest. 933 birds were evaluated for haemosporidian infections. We found a high parasitism prevalence (51.2%), which was correlated with phylogenetic relatedness among avian species. Prevalence varied drastically among the 20 well-sampled species, ranging from 0 to 70%. Seasonality was the main factor associated with infections, but how this abiotic condition influenced parasite prevalence varied according to the host-parasite system. Plasmodium prevalence increased during the rainy season and, after excluding the large sample size of Columbiformes (n = 462/933), Plasmodium infection rate was maintained high in the wet season and showed a negative association with host body mass. No association was found between non-Columbiform bird prevalence and seasonality or body mass when evaluating both Plasmodium and Haemoproteus or only Haemoproteus infections. Parasite community was composed of 32 lineages including 7 new lineages. We evidenced that even dry domains can harbour a high prevalence and diversity of vector-borne parasites and pointed out seasonality as a ruling factor.

Abstract Background Although haemosporidian parasites may cause considerable health and economic problems in aviaries, there is limited understanding of the vectors transmitting them. Mosquito-borne Plasmodium species are responsible for the deaths of numerous exotic (= immunologically naïve) birds in zoos every year, while native birds are adapted to the parasites and largely protected by an effective immune response. Methods Mosquitoes were collected in bird/animal parks, wetlands and private gardens in various regions of Germany from 2020 to 2022. Females were pooled with up to 10 specimens according to taxon, location and date. Extracted DNA was screened for avian Haemosporida-specific mitochondrial rDNA using real-time polymerase chain reaction (PCR). Positive samples were amplified by a Plasmodium/Haemoproteus-specific nested PCR targeting the partial cytochrome b gene, followed by sequencing of the PCR product for species identification. Sequences were checked against GenBank and MalAvi databases. Results PCR of 2633 pools with 8834 female mosquitoes signalled infection with Plasmodium in 46 pools and with Haemoproteus in one pool. Further amplification and sequencing demonstrated the occurrence of Haemoproteus majoris lineage PARUS1 (n = 1) as well as several Plasmodium species and lineages, including Plasmodium relictum SGS1 (n = 16) and GRW11 (n = 1), P. matutinum LINN1 (n = 13), P. vaughani SYAT05 (n = 10), P. circumflexum TURDUS01 (n = 3), P. cathemerium PADOM02 (n = 1) and Plasmodium sp. SYBOR02 (n = 1) and PLOPRI01 (n = 1). The infections were detected in Culex pipiens sensu lato (n = 40), Culiseta morsitans/fumipennis (n = 6) and Aedes cinereus/geminus (n = 1). Conclusions Although the overall Plasmodium minimum infection rate (5.2) appears to be low, the results demonstrated not only the ongoing circulation of Plasmodium parasites in the German mosquito population, but also the occurrence of eight distinct Plasmodium lineages, with three of them (PADOM02, SYBOR02, PLOPRI01) being detected in Germany for the first time. This study highlights the importance of conducting mosquito-borne pathogen surveillance studies simultaneously targeting vectors and vertebrate hosts, as certain species may be detected more readily in their vectors than in their vertebrate hosts, and vice versa. Graphical Abstract

Abstract Culex (Barraudius) modestus (Ficalbi 1889) are found in temperate regions across Europe, Asia, and Northern Africa. These mosquitoes thrive during the summer and prefer to breed in permanent vegetative habitats such as rice paddies and marshes. Culex modestus feed on a wide range of bird species but are highly attracted to humans, which makes them a potential ‘bridge’ vector for enzootic pathogens. There is compelling evidence that Culex modestus is an efficient vector for West Nile virus, potentially capable of causing epidemics in humans and other mammals. This species is also a likely vector for Usutu virus, avian malaria (Plasmodium spp.), and parasitic heartworms (Dirofilaria spp.). Culex modestus can be morphologically identified at the larval and adult stages, and a distinctive phenotype of this species is their ability to overwinter. Despite the widespread establishment of this mosquito species and their role as vectors for human pathogens, we lack sufficient knowledge on this species to implement and evaluate targeted vector control measures. Since Culex modestus can be considered a potential public health threat, there is a need for a better understanding of this mosquito species. Graphical Abstract

Host range expansions are an important factor for shaping the community of associated symbiotic organisms. Birds, as a highly mobile group of animals, are of particular interest to study with respect to the diversity and the distribution of the organisms using them as hosts during such large-scale movements. The Isabelline wheatear (Oenanthe isabellina) is a species with a main breeding area in Asia, which has expanded west, towards the Balkans, since the middle of the last century. We collected feather mites and blood samples for haemosporidian parasites screening from adult and juvenile Isabelline wheatears from the westernmost edge of the species breeding area in western Bulgaria. The feather mite species Alaudicola rosickyi (Černy, 1963), previously found on other wheatear species, was found for the first time on Isabelline wheatears. One species of avian malaria parasites—Plasmodium relictum (Grassi and Feletti, 1891) (lineage SGS1)—represented the first record of a malaria parasite in this host species in Europe. Increasing the sampling in the South European populations of the species will shed light on blood parasite species diversity and will reveal if other feather mite species have followed their host during its breeding range expansion.

Abstract Background Mitochondrial respiration plays a central role in the survival of many eukaryotes, including apicomplexan parasites. A 479-bp fragment from the mitochondrial cytochrome b gene is widely used as a barcode to identify genetic lineages of avian malaria parasites Plasmodium and related haemosporidians. Here we looked for evidence of selection in the avian Plasmodium cyt b gene, using tests of selection and protein structure modeling. We also tested for the association between cyt b polymorphism and the host specificity of these parasites. Results Based on 1,089 lineages retrieved from the Malavi database, we found that the frequency of the most conserved amino acids in most sites was more than 90%, indicating that the protein diversity of the avian Plasmodium cyt b barcode was low. The exceptions were four amino acid sites that were highly polymorphic, though the substitutions had only slight functional impacts on the encoded proteins. The selection analyses revealed that avian Plasmodium cyt b was under strong purifying selection, and no positively selected sites were detected. Besides, lineages with a wide host range tend to share cyt b protein haplotypes. Conclusions Our research indicates that purifying selection is the dominant force in the evolution of the avian Plasmodium cyt b lineages and leads to its low diversity at the protein level. Host specificity may also play a role in shaping the low mitochondrial diversity in the evolution of avian malaria parasites. Our results highlight the importance of considering selection pressure on the cyt b barcode region and lay a foundation for further understanding the evolutionary pattern of mitochondrial genes in avian malaria.

Background and Aim: Blood parasite infections in poultry, such as Plasmodium, are a serious threat to the poultry industry due to their potential to cause economic losses. To date, there has been inadequate research on the morphological and molecular detection of the different Plasmodium species that infect poultry in Indonesia. Therefore, this study aimed to analyze the morphological and molecular characteristics of Plasmodium spp. and the several predisposing factors for Plasmodium infection in layer chickens from three districts of Yogyakarta, Indonesia. Materials and Methods: One hundred and five blood samples from layer chickens were collected from 13 farms located in three districts of Yogyakarta (Sleman, Bantul, and Kulon Progo) between September and November 2022. Blood samples were subjected to microscopic and polymerase chain reaction (PCR) analyses. Sequencing was performed using basic local alignment search tools to identify the nucleotide structure of cytochrome b. Phylogenetic analysis of Plasmodium was performed using the MEGA-X software. Results: Microscopic examination revealed that 17/105 positives (16.19%) were positive for blood parasite infection. Trophozoites, erythrocytic meronts, and microgametocytes of Plasmodium were found in blood samples. Based on the morphological examination, the species found in the samples was close to Plasmodium juxtanucleare. Polymerase chain reaction examination revealed that 21/60 samples were positive for Plasmodium (35%). The Plasmodium species identified from the sequenced samples were proven to be P. juxtanucleare. The P. juxtanucleare from Thailand was closely related to samples (99.64%–100%) with a genetic distance of 0%–1%. In addition, age, population, and cage type were not significantly associated with Plasmodium infection. Conclusion: Based on microscopic and PCR examinations, the Plasmodium species found in the three districts of Yogyakarta was P. juxtanucleare. The genetic distance between samples from the three districts of Yogyakarta was closely related (0%–1%) to P. juxtanucleare from Thailand and Japan. There was no correlation between Plasmodium infection and age, cage type, or population.