Raising the bar: genus-specific nested PCR improves detection and lineage identification of avian haemosporidian parasites.

Avian haemosporidian parasites are useful model organisms to study the ecology and evolution of parasite-host interactions due to their global distribution and extensive biodiversity. Detection of these parasites has evolved from microscopic examination to PCR-based methods, with the mitochondrial cytochrome b gene serving as barcoding region. However, standard PCR protocols used for screening and identification purposes have limitations in detecting mixed infections and generating phylogenetically informative data due to short amplicon lengths. To address these issues, we developed a novel genus-specific nested PCR protocol targeting avian haemosporidian parasites. The protocol underwent rigorous testing utilizing a large dataset comprising blood samples from Malagasy birds of three distinct Passeriformes families. Furthermore, validation was done by examining smaller datasets in two other laboratories employing divergent master mixes and different bird species. Comparative analyses were conducted between the outcomes of the novel PCR protocol and those obtained through the widely used standard nested PCR method. The novel protocol enables specific identification of Plasmodium, Haemoproteus (Parahaemoproteus), and Leucocytozoon parasites. The analyses demonstrated comparable sensitivity to the standard nested PCR with notable improvements in detecting mixed infections. In addition, phylogenetic resolution is improved by amplification of longer fragments, leading to a better understanding of the haemosporidian biodiversity and evolution. Overall, the novel protocol represents a valuable addition to avian haemosporidian detection methodologies, facilitating comprehensive studies on parasite ecology, epidemiology, and evolution.

Morphological and molecular characterization of Eimeria haematopusi n. sp. (Apicomplexa: Eimeriidae) in an Australian Pied Oystercatcher (Haematopus longirostris) (Aves: Charadriiformes).

A novel Eimeria Schneider, 1875 species is described from an Australian pied oystercatcher Haematopus longirostris Vieillot, in Western Australia. The pied oystercatcher was admitted to the Kanyana Wildlife Rehabilitation Centre (KWRC), Perth, Western Australia in a poor body condition, abrasion to its right hock and signs of partial delamination to its lower beak. Investigation into potential medical causes resulted in a faecal sample being collected and screened for gastrointestinal parasites. Unsporulated coccidian oocysts were initially observed in the faeces and identified as Eimeria upon sporulation. The sporulated oocysts (n = 20) are ellipsoidal, 20-21 × 12-13 µm in shape and have thick bi-layered walls which are c.2/3 of the total thickness. Micropyle is present, robust and protruding, and occasionally has a rounded polar body attached to the micropyle. Within the oocyst, a residuum, in addition, two to five polar granules are present. There are four ellipsoidal sporocysts 9-11 × 5-6 µm with flattened to half-moon shaped Stieda bodies. Sub-Stieda body and para-Stieda body are absent. The sporocysts contain sporocyst residuums composed of a few spherules scattered among the sporozoites. Within the sporozoites, anterior and posterior refractile bodies are present, but the nucleus is indiscernible. To further characterise the novel Eimeria species from H. longirostris, molecular analysis was conducted at the 18S ribosomal RNA locus, using PCR amplification and cloning. Two cloned sequences from the novel Eimeria were compared with those from other Eimeria spp. with the highest genetic similarity of 97.6% and 97.2% from Clone 1 and 2, respectively with Eimeria reichenowi (AB544308) from a hooded crane (Grus monacha Temminck) in Japan. Both sequences grouped in a clade with the Eimeria spp. isolated from wetland birds, which include Eimeria paludosa (KJ767187) from a dusky moorhen (Gallinula tenebrosa Gould) in Western Australia, Eimeria reichenowi (AB544308) and Eimeria gruis (AB544336) both from hooded cranes. Based on the morphological and molecular data, this Eimeria sp. is a new species of coccidian parasite and is named Eimeria haematopusi n. sp. after its host H. longirostris.

Ticks (Acari: Ixodidae) on resident and migratory wild birds in Orinoquia region, Colombia.

Several species of hard ticks, including those of the genera Ixodes, Haemaphysalis, Amblyomma, and Rhipicephalus, are of medical and veterinary importance and have been reported in association with Neotropical wild birds. Colombia, known for its great bird diversity, has 57 confirmed tick species. However, there are few studies on the association between wild birds and ticks in Colombia. The Orinoquia region, a migratory center in Colombia, provides a unique opportunity to study wild bird-tick associations and their implications for tick-borne disease dynamics. Our study, conducted between October and December 2021, aimed to identify hard ticks infesting resident and migratory wild birds in the department of Arauca and to assess the presence of bacteria from the genera Anaplasma, Borrelia, Ehrlichia, Rickettsia, and piroplasms. A total of 383 birds were examined, of which 21 were infested. We collected 147 ticks, including Amblyomma dissimile (larvae), Amblyomma longirostre (nymphs), Amblyomma mixtum (adults), and Amblyomma nodosum (larvae and nymphs). We did not detect bacterial DNA in the tested ticks; however, piroplasm DNA was detected in ticks from three of the infested birds. Of the 21 bird-tick associations, six are new to the Americas, and interesting documentation of piroplasm DNA in A. longirostre, A. nodosum, and A. dissimile ticks from wild birds in the region. This study provides valuable insights into the ticks associated with wild birds and their role in the dispersal of ticks and pathogens in Colombia, enhancing our understanding of tick life cycles and tick-borne disease dynamics.

INFLUENCE OF GROOMING ON PERMANENT ARTHROPOD ASSOCIATES OF BIRDS: CATTLE EGRETS, LICE, AND MITES.

Birds have a diverse community of "permanent" arthropods that complete their entire life cycle on the body of the host. Because some of these arthropods are parasites that reduce host fitness, birds control them by grooming, which consists of preening with the beak and scratching with the feet. Although preening is the primary component of grooming, scratching is essential for controlling arthropods on the head and neck, which cannot be preened. Several unrelated groups of birds have evolved comb-like pectinate claws on the middle toenail of each foot. We tested the role of these claws in the control of arthropods by experimentally removing teeth from the claws of captive western cattle egrets (Bubulcus ibis) infested with chewing lice (Insecta: Phthiraptera), feather mites (Acari: Sarcoptiformes), and nasal mites (Acari: Mesostigmata). After a period of 4 mo, we compared the abundance of arthropods on experimental birds to that of control birds with intact teeth. We used video to quantify the grooming rates of the captive birds, which groomed twice as much as wild birds. Experimental and control birds did not differ significantly in grooming time. Both groups virtually eradicated the chewing lice, but not feather mites or nasal mites. We found no support for the hypothesis that pectinate claws increase the efficiency of arthropod control by grooming. Experiments with wild birds are needed to test the hypothesis further under conditions in which birds devote less time to grooming.

Selection of suitable reference genes for normalization of RT-qPCR in three tissues of Northern bobwhite (Colinus virginianus) infected with eyeworm (Oxyspirura petrowi).

BACKGROUND: The Northern bobwhite (Colinus virginianus) is an economically important, and popular game bird in North America. Northern bobwhites have experiencing declines of > 3.5% annually in recent decades due to several factors. The eyeworm Oxyspirura petrowi is a nematode parasite frequently found in the eyes of bobwhites. Although reported frequently in wild bobwhites, there is no research to understand the host-parasite mechanism. Hence, it is important to investigate mechanisms of eyeworm invasion and immune modulation in bobwhite. Cytokine gene expression using RT-PCR is widely used to identify the innate immune response of a host to an infection. METHODOLOGY: In this study, we evaluated ten reference genes (HMBS, RPL19, RPL32, RPS7, RPS8, TATA, SDHA, YWHAZ, GAPDH, and ACTB) for their stability across three tissues (liver, spleen, and caecal tonsils) of control and O. petrowi infected Northern bobwhites. Primer efficiency and reference genes stability were assessed using GeNorm, NormFinder, and BestKeeper. RESULTS: Expression of these reference genes with respect to O. petrowi infection in bobwhites showed RPL32 and HMBS were the most stable genes in the liver, HMBS and SDHA were the most stable genes in the spleen, and HMBS and YWHAZ were equally stable reference genes in the caecal tonsils. CONCLUSION: Based on the geometric mean of all three analyses, our results indicate that the combination of RPL32 and HMBS for the liver, HMBS and SDHA for the spleen, and YWHAZ and HMBS for caecal tonsils might be used as reference genes for normalization in gene expression investigations on Northern bobwhites.

First report of Sarcocystis falcatula in naturally infected Razorbill auks (Alca torda) collected in Tunisian Mediterranean Sea shores.

Sarcocystis spp. are apicomplexan cyst-forming parasites that can infect numerous vertebrates, including birds. Sarcosporidiosis infection was investigated in three muscles (breast, right and left thigh muscle) and one organ (heart) of four Razorbill auks (Alca torda) stranded between November and December 2022 on the shores of the Mediterranean Sea in Nabeul and Bizerte governorates, Northern Tunisia. Two of the four tested A. torda were PCR positive for 18S rRNA Sarcocystis spp. gene. Among the examined 16 muscles/organs, only one breast and one right thigh were Sarcocystis spp. PCR-positive (12.5% ± 8.3, 2/16). Our results showed a relatively high molecular prevalence of Sarcocystis spp. in Razorbill auks (A. torda). Sarcocystis spp. sequence described in the present study (GenBank number: OR516818) showed 99.56-100% identity to Sarcocystis falcatula. In conclusion, our results confirmed the infection of Razorbill auks (A. torda) by S. falcatula. Further research is needed on different migratory seabirds' species in order to identify other Sarcocystis species.

Molecular detection of Toxoplasma gondii and Neospora caninum in seabirds collected along the coast of Santa Catarina, Brazil.

Toxoplasma gondii and Neospora caninum are two closely related protozoans that infect a wide range of animals, including birds. However, the occurrence of N. caninum and T. gondii in seabirds is unknown. Therefore, this study aimed to determine the presence of T. gondii and N. caninum DNA in tissue samples of seabirds. Tissue samples of the pectoral muscles, heart, and brain were collected from 47 birds along the coastline of Santa Catarina State, SC, Brazil. The DNA was extracted from the tissues and screened using nested-PCR (nPCR) targeting internal transcribed spacer 1 (ITS1). T. gondii DNA was detected in tissues from seven seabirds (7/47, 14.8%), kelp gull (Larus dominicanus) (5/21), and Manx shearwater (Puffinus puffinus) (2/8). N. caninum DNA was detected in tissues of nine seabirds (9/47, 19.1%), the kelp gull (L. dominicanus) (4/21), Manx shearwater (P. puffinus) (2/8), neotropic cormorant (Phalacrocorax brasilianus) (1/4), brown booby (Sula leucogaster) (1/5), and white-chinned petrel (Procellaria aequinoctialis) (1/1); however, no co-infection was observed. In conclusion, this study showed the circulation of N. caninum and T. gondii in seabirds along the coastline of Santa Catarina State. Further studies are required to clarify the role of these birds in the epidemiology of neosporosis and toxoplasmosis.

High prevalence of haemosporidian parasites in Eurasian jays.

Avian haemosporidians are vector-borne parasites, infecting a great variety of birds. The order Passeriformes has the highest average infection probability; nevertheless, some common species of Passeriformes have been rather poorly studied. We investigated haemosporidians in one such species, the Eurasian jay Garrulus glandarius (Corvidae), from a forest population in Hesse, Central Germany. All individuals were infected with at least one haemosporidian genus (overall prevalence: 100%). The most common infection pattern was a mixed Haemoproteus and Leucocytozoon infection, whereas no Plasmodium infection was detected. Results on lineage diversity indicate a rather pronounced host-specificity of Haemoproteus and Leucocytozoon lineages infecting birds of the family Corvidae.

Coinfection of slime feather duster worms (Annelida, Myxicola) by different gregarine apicomplexans (Selenidium) and astome ciliates reflects spatial niche partitioning and host specificity.

Individual organisms can host multiple species of parasites (or symbionts), and one species of parasite can infect different host species, creating complex interactions among multiple hosts and parasites. When multiple parasite species coexist in a host, they may compete or use strategies, such as spatial niche partitioning, to reduce competition. Here, we present a host­symbiont system with two species of Selenidium (Apicomplexa, Gregarinida) and one species of astome ciliate co-infecting two different species of slime feather duster worms (Annelida, Sabellidae, Myxicola) living in neighbouring habitats. We examined the morphology of the endosymbionts with light and scanning electron microscopy (SEM) and inferred their phylogenetic interrelationships using small subunit (SSU) rDNA sequences. In the host 'Myxicola sp. Quadra', we found two distinct species of Selenidium; S. cf. mesnili exclusively inhabited the foregut, and S. elongatum n. sp. inhabited the mid to hindgut, reflecting spatial niche partitioning. Selenidium elongatum n. sp. was also present in the host M. aesthetica, which harboured the astome ciliate Pennarella elegantia n. gen. et sp. Selenidium cf. mesnili and P. elegantia n. gen. et sp. were absent in the other host species, indicating host specificity. This system offers an intriguing opportunity to explore diverse aspects of host­endosymbiont interactions and competition among endosymbionts.

First Report of Intestinal Cestode Infection in a Berylline Hummingbird (Saucerottia beryllina) in Mexico.

BACKGROUND: Up to now, five cestode species have been reported infecting five hummingbird species. To date, there have been no reports of cestode infections in hummingbirds in Mexico. METHODS: A Berylline hummingbird (Saucerottia beryllina) was found dead in a backyard at Toluca City, Mexico, and a window collision was assumed as the cause of death. The bird was preserved in 10% neutral buffered formalin for routine histological examination. RESULTS: At the histological study, liver parenchymal disruption was observed. This lesion could be the result of the assume collision. No lesions were observed in other tissues examined. Conspicuous cestode structures were observed in the lumen of the small intestine. Structure of cestodes, as revealed from histological sections, suggests their position in the genus Anonchotaenia Cohn, 1900 (family Paruterinidae). CONCLUSION: This is the first report of intestinal cestodosis in a Berylline hummingbird (S. beryllina) in Mexico.

Prevalence and molecular characterization of Trichomonas gallinae from pigeons in Anhui, China.

Trichomonas gallinae, a protozoan parasite causing avian trichomonosis, exhibits a widespread global prevalence. It primarily affects the upper digestive tract of birds and has resulted in significant ecological problems worldwide. This study aimed to investigate the prevalence and genotypes of T. gallinae in Anhui Province, China. A total of 1612 oropharyngeal swab samples were collected from pigeon farms in Anhui Province to determine the prevalence of T. gallinae infection. The results revealed 565 (35.1%) positive samples of T. gallinae. Significant differences in infection rates were observed among different regions and age groups. Furthermore, the ITS1/5.8 S/ITS2 region was amplified, sequenced, and subjected to phylogenetic analysis. Genotypes A and B of T. gallinae were identified, and genotype B was the dominant genotype in Anhui Province. This is the first report on the prevalence and molecular characterization of T. gallinae in Anhui Province, China. Additionally, we integrated reports on the prevalence and genotype of T. gallinae in relevant provinces in China.

Brain parasites and misorientation of migratory birds.

Haemoproteus blood parasites of birds are thought to be relatively benign. Recent findings show that infections may develop in the brain of birds, possibly distorting their orientation sense. Misdirected migration may lead migrants outside their range where they are recognized as vagrants and can transmit parasites to novel hosts.

Molecular characterization of Haemoproteus enucleator with emphasis on the host and geographic distribution.

Haemoproteus species (Haemosporida, Haemoproteidae) are cosmopolitan and highly diverse blood parasites of birds that have been neglected in avian medicine. However, recent discoveries based on molecular diagnostic markers show that these pathogens often cause marked damage to various internal organs due to exo-erythrocytic development, sometimes resulting in severe and even lethal avian haemoproteosis, including cerebral pathologies. Molecular markers are essential for haemoproteosis diagnostics, but the data is limited, particularly for parasites transmitted in tropical ecosystems. This study combined microscopic and molecular approaches to characterize Haemoproteus enucleator morphologically and molecularly. Blood samples were collected from the African pygmy kingfisher Ispidina picta in Cameroon, and the parasite was identified using morphological characters of gametocytes. The analysis of partial cytochrome b sequences (cytb) identified a new Haemoproteus lineage (hISPIC03), which was linked to the morphospecies H. enucleator. Illustrations of blood stages were provided and the phylogenetic analysis showed that the new lineage clustered with five other closely related lineages belonging to the same morphospecies (hALCLEU01, hALCLEU02, hALCLEU03, hISPIC01, and hALCQUA01), with a maximum genetic distance between these lineages of 1.5 % (7 bp difference) in the 478 bp cytb sequences. DNA haplotype network was developed and identified geographic and host distribution of all lineages belonging to H. enucleator group. These lineages were almost exclusively detected in African kingfishers from Gabon, Cameroon, South Africa, and Botswana. This study developed the molecular characterization of H. enucleator and provides opportunities for diagnostics of this pathogen at all stages of its life cycle, which remains undescribed in all its closely related lineages.

Urbanization correlates with the prevalence and richness of blood parasites in Eurasian Blackbirds (Turdus merula).

Urbanization is increasing worldwide, producing severe environmental impacts. Biodiversity is affected by the expansion of cities, with many species being unable to cope with the different human-induced stressors present in these landscapes. However, this knowledge is mainly based on research from taxa such as plants or vertebrates, while other organisms like protozoa have been less studied in this context. The impact of urbanization on the transmission of vector-borne pathogens in wildlife is still unclear despite its relevance for animal and human health. Here, we investigated whether cities are associated with changes in the prevalence and richness of lineages of three vector-borne protozoans (Plasmodium, Haemoproteus and Leucocytozoon) in Eurasian blackbirds (Turdus merula) from multiple urban and forest areas in Europe. Our results show important species-specific differences between these two habitat types. We found a significant lower prevalence of Leucocytozoon in urban birds compared to forest birds, but no differences for Plasmodium and Haemoproteus. Furthermore, the richness of parasite lineages in European cities was higher for Plasmodium but lower for Leucocytozoon than in forests. We also found one Plasmodium lineage exclusively from cities while another of Leucocytozoon was only found in forests suggesting a certain level of habitat specialization for these protozoan vectors. Overall, our findings show that cities provide contrasting opportunities for the transmission of different vector-borne pathogens and generate new scenarios for the interactions between hosts, vectors and parasites.

Surveillance for Selected Pathogens and Parasites of Northern Bobwhite (Colinus virginianus) from Western Oklahoma, USA, 2018-20.

The Northern Bobwhite (Colinus virginianus) has been undergoing a range-wide population decline. Potential causes for declines across its historic range have been investigated for decades and include habitat loss and fragmentation and a variety of parasitic and infectious diseases. Although there have been studies on bobwhite ecology in Oklahoma, USA, relatively little is known about parasites and pathogens in the region. We evaluated the health of free-ranging bobwhites from nine sites in western Oklahoma. From 2018 to 2020, 206 bobwhites were evaluated for gross and microscopic lesions and tested for selected pathogens. In general, bobwhites were in good nutritional condition with ample muscle mass and fat stores. No significant gross lesions were observed in any bobwhite and no significant histologic lesions were detected in a subset. There was no evidence of infection with or exposure to reticuloendotheliosis virus, West Nile virus, respiratory Mycoplasmataceae species, Pasteurella multocida, intestinal Eimeria spp., or oral Trichomonas spp. Several pathogens of potential concern were detected, including avian adenovirus (8.6%), Toxoplasma gondii (2.3%), and haemosporidians (a Haemoproteus sp. (1.5%), Leucocytozoon schoutedeni (1.5%), and Plasmodium homopolare haplotype 2 [lineage LAIRI01; 3.6%]). Physaloptera sp. (12%) and Sarcocystis sp. (1%) were detected in the breast muscle. Low intraspecific genetic diversity was noted for Physaloptera sp., and sequences were most similar to Physaloptera sequences from bobwhites and grasshoppers (Orthoptera) in Texas. Low intensities of chewing lice, chiggers, and ticks were observed. A subset of bobwhites had evidence of exposure to selected toxicants and heavy metals; a small number had low levels of iron, manganese, zinc, molybdenum, and copper, which were not considered diagnostically relevant. In general, bobwhites from western Oklahoma appeared to be in good health with a low diversity of pathogens detected, but future work is needed to understand potentially changing disease risks for this population.

Novel phylogenetic clade of avian Haemoproteus parasites (Haemosporida, Haemoproteidae) from Accipitridae raptors, with description of a new Haemoproteus species.

Avian haemosporidian parasites (order Haemosporida, phylum Apicomplexa) are blood and tissue parasites transmitted by blood-sucking dipteran insects. Three genera (Plasmodium, Haemoproteus and Leucocytozoon) have been most often found in birds, with over 270 species described and named in avian hosts based mainly on the morphological characters of blood stages. A broad diversity of Haemoproteus parasites remains to be identified and characterized morphologically and molecularly, especially those infecting birds of prey, an underrepresented bird group in haemosporidian parasite studies. The aim of this study was to investigate and identify Haemoproteus parasites from a large sample comprising accipitriform raptors of 16 species combining morphological and new molecular protocols targeting the cytb genes of this parasite group. This study provides morphological descriptions and molecular characterizations of two Haemoproteus species, H. multivacuolatus n. sp. and H. nisi Peirce and Marquiss, 1983. Haemoproteus parasites of this group were so far found in accipitriform raptors only and might be classified into a separate subgenus or even genus. Cytb sequences of these parasites diverge by more than 15% from those of all others known avian haemosporidian genera and form a unique phylogenetic clade. This study underlines the importance of developing new diagnostic tools to detect molecularly highly divergent parasites that might be undetectable by commonly used conventional tools.

Title: Nouveau clade phylogénétique de parasites de rapaces Accipitridae du genre Haemoproteus (Haemosporida, Haemoproteidae), avec description d'une nouvelle espèce d'Haemoproteus. Abstract: Les parasites hémosporidies aviaires (ordre Haemosporida, phylum Apicomplexa) sont des parasites sanguins et tissulaires transmis par des insectes diptères hématophages. Trois genres (Plasmodium, Haemoproteus et Leucocytozoon) ont été le plus souvent trouvés chez les oiseaux, avec plus de 270 espèces décrites et nommées chez les hôtes aviaires en fonction principalement des caractères morphologiques des stades sanguins. Une grande diversité des Haemoproteus reste à identifier et à caractériser morphologiquement et génétiquement, en particulier ceux qui infectent les oiseaux de proie, un groupe d'oiseaux sous-représenté dans les études sur les hémosporidies. Le but de cette étude était d'étudier et d'identifier les Haemoproteus à partir d'un large échantillon comprenant des rapaces accipitriformes de 16 espèces, en combinant des protocoles morphologiques et de nouveaux protocoles moléculaires ciblant les gènes cytb de ce groupe de parasites. Cette étude fournit des descriptions morphologiques et des caractérisations moléculaires de deux espèces d'Haemoproteus, H. multivacuolatus n. sp. et H. nisi Peirce and Marquiss, 1983. Les Haemoproteus de ce groupe n'ont jusqu'à présent été trouvés que chez les rapaces accipitriformes et pourraient être classés dans un sous-genre ou même un genre distinct. Les séquences cytb de ces parasites divergent de plus de 15 % de celles de tous les autres genres d'hémosporidies aviaires connus et forment un clade phylogénétique unique. Cette étude souligne l'importance de développer de nouveaux outils de diagnostic pour détecter des parasites moléculairement très divergents qui pourraient être indétectables par les outils conventionnels couramment utilisés.

Chewing lice of Bearded Reedling (Panurus biarmicus) and diversity of louse-host associations of birds in reed beds in Slovakia.

A total of 1,621 wild birds representing 34 species were examined for chewing lice in reed beds in southwestern Slovakia during the pre-breeding migration 2008-2009 and 2016-2019. A total of 377 (23.3%) birds representing 15 species were parasitized by 26 species of chewing lice of 12 genera. Dominant genera were Penenirmus (with dominance 32.6%) and Menacanthus (29.4%), followed by Brueelia (12.6%), Acronirmus (10.8%), Philopterus (7.7%), and Myrsidea (4.2%). We evaluated 33 host-louse associations including both 1) host-generalist, parasitizing more than one host species and host-specific lice, occurring only on a single host species, and 2) lice species with large range geographic distribution, reported across the range of the distribution of their hosts and lice species with only occasional records from a limited area within the range of their hosts. The Bearded Reedling, Panurus biarmicus (Linnaeus, 1758), was parasitized by two species of chewing lice, Menacanthus brelihi Balát, 1981 and Penenirmus visendus (Zlotorzycka, 1964), with conspicuously different prevalences (5.6% vs. 58.2%, respectively; n = 251). New material enabled us to redescribe both species of lice: the first one is resurrected from previous synonymy as a valid species. A fragment of the mitochondrial cytochrome oxidase I gene was sequenced from these two species in order to assess their relative phylogenetic position within their genera. Our study demonstrates the importance of an adequate identification of parasites, especially on rarely examined and endangered hosts.

Title: Mallophages de la Panure à moustaches (Panurus biarmicus) et diversité des associations mallophages-hôtes des oiseaux dans les roselières en Slovaquie. Abstract: Au total, 1 621 oiseaux sauvages représentant 34 espèces ont été examinés à la recherche de mallophages dans les roselières du sud-ouest de la Slovaquie au cours de la migration de pré-reproduction 2008­2009 et 2016­2019. Parmi ceux-ci, 377 oiseaux (23,3 %), représentant 15 espèces, étaient parasités par 26 espèces de mallophages de 12 genres. Les genres dominants étaient Penenirmus (avec une dominance de 32,6 %) et Menacanthus (29,4 %), suivis de Brueelia (12,6 %), Acronirmus (10,8 %), Philopterus (7,7 %) et Myrsidea (4,2 %). Nous avons évalué 33 associations mallophage-hôte comprenant à la fois 1) des espèces de mallophages généralistes, parasitant plus d'une espèce hôte, et des mallophages spécifiques, présents uniquement sur une seule espèce hôte et 2) des espèces de mallophages ayant une large répartition géographique, signalées à travers l'étendue de la répartition de leurs hôtes, et des espèces de mallophages avec seulement des observations occasionnelles dans une zone limitée à l'intérieur de l'aire de répartition de leurs hôtes. La Panure à moustaches, Panurus biarmicus (Linnaeus, 1758), était parasitée par deux espèces de mallophages, Menacanthus brelihi Balát, 1981 et Penenirmus visendus (Zlotorzycka, 1964), avec des prévalences nettement différentes (respectivement 5,6 % et 58,2 %, n = 251). Du nouveau matériel nous a permis de redécrire les deux espèces de mallophages, la première étant ressuscitée de la synonymie précédente en tant qu'espèce valide. Un fragment du gène mitochondrial de la cytochrome oxydase I a été séquencé à partir de ces deux espèces afin d'évaluer leur position phylogénétique relative au sein de leurs genres. Notre étude démontre l'importance d'une identification adéquate des parasites, en particulier sur les hôtes rarement examinés et menacés.

IMPLICATIONS OF TREATING PARASITIC INFECTION IN NORTHERN BOBWHITE (COLINUS VIRGINIANUS) ON OVERALL HELMINTH LIFE CYCLE.

This study was conducted to determine how reducing the parasite burden in a definitive host may affect the prevalence in intermediate hosts. Here we used the eyeworm Oxyspirura petrowi and cecal worm Aulonocephalus pennula as model species. Northern bobwhite quail (Colinus virginianus) were provided an anthelmintic medicated feed in wild systems because of convincing evidence that these parasites were suppressing their populations. Eyeworm and cecal worm prevalence were measured in Orthopterans, which act as intermediate hosts, using polymerase chain reaction. Individuals were collected from a control site, a site treated for 2 yr, and a site treated for 5 yr. Orthopteran community composition was significantly different among the sites; however, an interaction between subfamily and site was not significant for the eyeworm. There was a significant reduction in eyeworm-infected Orthopterans on the 5-yr site compared with the other 2, suggesting that treatment of a definitive host may indeed affect the prevalence of eyeworms during other life-cycle stages. There was an interaction between the Orthopteran subfamily and the site for the cecal worm, so results were analyzed within each subfamily. A significant reduction in the prevalence of cecal worms was only found in the Cyrtacanthacridinae subfamily on the 5-yr site when compared with the other sites. However, the greatest prevalence in the Gomphocerinae and Oedipodinae subfamilies across all 3 sites was 4.1%. This indicates an unknown degree of cecal worm host specificity. Therefore, conclusions could not be made through the simple assessment of prevalence.

Molecular Survey of Haemosporidian Parasites in Procellariiformes Sampled in Southern Brazil, 2013-22.

The order Procellariiformes includes several species of seabirds that perform long-distance migrations crossing all the oceans. These movements may contribute to the dispersal and exchange of hemoparasites, such as haemosporidians. There is a lack of studies regarding the order Haemosporida in Procellariiformes, and, to date, only the genus Plasmodium has been reported. This survey investigated the occurrence of the three genera of haemosporidians, Plasmodium, Haemoproteus, and Leucocytozoon, in samples collected between 2013 and 2022 from 95 individuals of 14 species of Procellariiformes from southern Brazil, including live animals in rehabilitation centers, individuals caught as incidental bycatch, and carcasses found along the coast. A total of 171 samples of blood and fragments of liver and spleen were analyzed, with extracted DNA being subjected to a nested PCR followed by phylogeny analysis. All animals were negative for Plasmodium spp. and Leucocytozoon spp., but one Black-browed Albatross (Thalassarche melanophris) and one Manx Shearwater (Puffinus puffinus) specimen were positive for Haemoproteus spp. The sequences obtained from positive seabirds did not show 100% similarity with other known lineages available in the MalAvi database and thus were probably novel lineages. However, one sequence clustered together with Haemoproteus noctuae, a parasite from Strigiformes, while the other was grouped with Haemoproteus columbae, which is classically related to Columbiformes. These results suggest that both positive animals may have become infected when beached or in rehabilitation centers by a spillover of vectors from local birds. This highlights the importance of surveillance of the health of Procellariiformes regarding the possibility of dissemination of new pathogens in different bird populations.

Co-infecting Haemoproteus species (Haemosporida, Apicomplexa) show different host tissue tropism during exo-erythrocytic development in Fringilla coelebs (Fringillidae).

Avian haemosporidians of the genera Plasmodium, Haemoproteus, and Leucocytozoon are common blood parasites in wild birds all over the world. Despite their importance as pathogens potentially compromising host fitness and health, little is known about the exo-erythrocytic development of these parasites, particularly during co-infections which predominate in wildlife. This study aimed to address this issue using Haemoproteus parasites of Fringilla coelebs, a common bird species of the Western Palearctic and host to a variety of haemosporidian parasite lineages. Blood and tissue samples of 20 F. coelebs, positive for haemosporidians by blood film microscopy, were analysed by PCR and sequencing to determine cytochrome b lineages of the parasites. Tissue sections were examined for exo-erythrocytic stages by histology and in situ hybridization applying genus-, species-, and lineage-specific probes which target the 18S rRNA of the parasites. In addition, laser microdissection of tissue stages was performed to identify parasite lineages. Combined molecular results of PCR, laser microdissection, and in situ hybridization showed a high rate of co-infections, with Haemoproteus lineages dominating. Exo-erythrocytic meronts of five Haemoproteus spp. were described for the first known time, including Haemoproteus magnus hCCF6, Haemoproteus fringillae hCCF3, Haemoproteus majoris hCCF5, Haemoproteus sp. hROFI1, and Haemoproteus sp. hCCF2. Merogonic stages were observed in the vascular system, presenting a formerly unknown mode of exo-erythrocytic development in Haemoproteus parasites. Meronts and megalomeronts of these species were distinct regarding their morphology and organ distribution, indicating species-specific patterns of merogony and different host tissue tropism. New pathological aspects of haemoproteosis were reported. Furthermore, phylogenetic analysis of Haemoproteus spp. with regard to their exo-erythrocytic stages points towards separation of non-megalomeront-forming species from megalomeront-forming species, calling for further studies on exo-erythrocytic development of haemosporidian parasites to explore the phylogenetic character of this trait.