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Background Eukaryotic pathogens, including Cryptosporidium , Giardia and Enterocytozoon , have been implicated in neonatal diarrhoea, leading to marked morbidity and mortality in the alpaca ( Vicugna pacos ) and llama ( Lama glama ) around the world. Australia has the largest population of alpacas outside of South America, but very little is known about these pathogens in alpaca populations in this country. Here, we undertook the first molecular epidemiological survey of Cryptosporidium , Giardia and Enterocytozoon in V. pacos in Australia. Methods A cross-sectional survey of 81 herds, comprising alpacas of 6 weeks to 26 years of age, were sampled from the six Australian states (Queensland, New South Wales, Victoria, South Australia, Tasmania and Western Australia) across the four seasons. PCR-based sequencing was employed, utilising genetic markers in the small subunit of the nuclear ribosomal RNA ( SSU ) and 60-kilodalton glycoprotein ( gp60 ) genes for Cryptosporidium , triose-phosphate isomerase ( tpi ) gene for Giardia duodenalis and the internal transcribed spacer region ( ITS ) for Enterocytozoon bieneusi . Results PCR-based analyses of 81 faecal DNA samples representing 1421 alpaca individuals detected Cryptosporidium , Giardia and/or Enterocytozoon on 15 farms in New South Wales, Victoria and South Australia, equating to 18.5% of all samples/herds tested. Cryptosporidium was detected on three (3.7%) farms, G. duodenalis on six (7.4%) and E. bieneusi on eight (9.9%) in two or all of these three states, but not in Queensland, Tasmania or Western Australia . Molecular analyses of selected faecal DNA samples from individual alpacas for Cryptosporidium , Giardia and/or Enterocytozoon consistently showed that alpacas of ≤ 6 months of age harboured these pathogens. Conclusions This first molecular investigation of Cryptosporidium , Giardia and Enterocytozoon in alpaca subpopulations in Australia has identified species and genotypes that are of likely importance as primary pathogens of alpacas, particularly young crias, and some genotypes with zoonotic potential. Although the prevalence established here in the alpaca subpopulations studied is low, the present findings suggest that crias are likely reservoirs of infections to susceptible alpacas and/or humans. Future studies should focus on investigating pre-weaned and post-weaned crias, and on exploring transmission patterns to establish what role particular genotypes play in neonatal or perinatal diarrhoea in alpacas and in zoonotic diseases in different states of Australia.

Background Gastrointestinal nematodes (GINs) can cause significant economic losses in alpacas due to lowered production of fibre and meat. Although no anthelmintics are registered for use in alpacas, various classes of anthelmintics are frequently used to control parasitic gastroenteritis in alpacas in Australia and other countries. Very little is known about the current worm control practices as well as the efficacy of anthelmintics used against common GINs of alpacas. This study aimed to assess the existing worm control practices used by Australian alpaca farmers and to quantify the efficacy of commonly used anthelmintics against GINs of alpacas. Methods An online questionnaire survey was conducted to assess current worm control practices on 97 Australian alpaca farms, with an emphasis on the use of anthelmintics. Of this group of 97 alpaca farms, 20 were selected to assess the efficacy of eight anthelmintics and/or their combinations (closantel, fenbendazole ivermectin, monepantel, moxidectin and a combination of levamisole, closantel, albendazole, abamectin) using the faecal egg count reduction test (FECRT). A multiplexed-tandem PCR (MT-PCR) was used to identify the prevalent nematode genera/species. Results The response rate for the questionnaire was 94% (91/97). Almost half of the respondents kept alpacas with sheep and cattle, and 26% of respondents allowed alpacas to co-graze with these ruminants. Although only 63% respondents perceived worms to be an important health concern for alpacas, the majority of respondents (89%) used anthelmintics to control GINs of alpacas. The commonly used anthelmintics were macrocyclic lactones, monepantel, benzimidazoles, levamisole, closantel and their combinations, and they were typically administered at the dose rate recommended for sheep. The FECRT results showed that a combination of levamisole, closantel, albendazole and abamectin was the most effective dewormer followed by single drugs, including monepantel, moxidectin, closantel, fenbendazole and ivermectin. Haemonchus spp. were the most commonly resistant nematodes followed by Trichostrongylus spp., Camelostrongylus mentulatus , Ostertagia ostertagi and Cooperia spp. Conclusions This is the first study aimed at assessing worm control practices and efficacy of commonly used anthelmintics in alpacas in Australia. Our findings document the extent of anthelmintics resistance on Australian alpaca farms and identify those anthelmintics that are still effective against GINs of alpacas.

This study aimed to compare the FECPAK G2 and the McMaster techniques for counting of gastrointestinal nematode eggs in the faeces of alpacas using two floatation solutions (saturated sodium chloride and sucrose solutions). Faecal eggs counts from both techniques were compared using the Lin’s concordance correlation coefficient and Bland and Altman statistics. Results showed moderate to good agreement between the two methods, with better agreement achieved when saturated sugar is used as a floatation fluid, particularly when faecal egg counts are less than 1000 eggs per gram of faeces. To the best of our knowledge this is the first study to assess agreement of measurements between McMaster and FECPAK G2 methods for estimating faecal eggs in South American camelids.

Sarcocystis spp. are intracellular coccidian parasites which infect domestic and wild animals and birds, resulting in considerable economic losses in production animals, and public health concerns worldwide. Sarcocystis spp. have an indirect life cycle where wild and/or domestic canine species primarily serve as definitive hosts and several domestic and wild animals (such as camels) act as intermediate hosts. In Northern Africa, the Middle East, Central Asia and China, camel meat is preferred due to cultural and religious traditions as well as its lower cholesterol/fat content than other red meat. However, camel meat quality could be downgraded by the presence of sarcocysts. To date, two Sarcocystis spp. have been reported from camels, including Sarcocystis ippeni (forms microscopic sarcocysts) and Sarcocystis cameli (forms both macroscopic and microscopic sarcocysts). Sarcocystosis is usually asymptomatic, though significant pathogenic effects have also been reported in camels. Despite the high occurrence of sarcocystosis in camels, little is known about various aspects of the disease in these animals. This paper provides a comprehensive review of the existing knowledge on the taxonomy, pathogenesis, epidemiology and diagnosis of Sarcocystis spp. infecting camels and it also highlights areas for further research that could enhance our understanding about sarcocystosis in camels.

The adequate transfer of passive immunity is a critical factor in neonatal development and survivability. Although well documented in the dairy and equine industries, the recognition of inadequate immunoglobulin transfer on-farm and its impact on the ability of alpaca cria to thrive is largely unknown. Colostrum samples were collected from female alpaca within 24 h of parturition by the owners and whole blood collected from cria by the investigators between 1 and 7 days of age. Direct IgG concentration of milk and serum was determined using radial immunodiffusion assay (RID) and was indirectly estimated using optical and digital Brix refractometry for total solids and clinical refractometry for total serum protein. There was a strong correlation between optical and digital Brix refractometry, and colostral IgG concentration determined by RID. There was a moderate correlation between serum IgG concentration determined by RID and total serum protein in crias. Optical and digital Brix refractometry for colostral IgG estimation and total serum protein for serum IgG estimation are reliable, accurate and easy-to-use tools that can be used on-farm by trained, competent technicians to assess a failure of passive transfer in alpacas. A pilot study at one property only was performed, due to COVID-19 travel restriction interference. Further research is required to determine the reference intervals for these tools to be practical.

This study aimed to compare the FECPAK and the McMaster techniques for counting of gastrointestinal nematode eggs in the faeces of alpacas using two floatation solutions (saturated sodium chloride and sucrose solutions). Faecal eggs counts from both techniques were compared using the Lin's concordance correlation coefficient and Bland and Altman statistics. Results showed moderate to good agreement between the two methods, with better agreement achieved when saturated sugar is used as a floatation fluid, particularly when faecal egg counts are less than 1000 eggs per gram of faeces. To the best of our knowledge this is the first study to assess agreement of measurements between McMaster and FECPAK methods for estimating faecal eggs in South American camelids.

Background The gibberellin (GA) pathway plays a central role in the regulation of plant development, with the 2-oxoglutarate-dependent dioxygenases (2-ODDs: GA20ox, GA3ox, GA2ox) that catalyse the later steps in the biosynthetic pathway of particularly importance in regulating bioactive GA levels. Although GA has important impacts on crop yield and quality, our understanding of the regulation of GA biosynthesis during wheat and barley development remains limited. In this study we identified or assembled genes encoding the GA 2-ODDs of wheat, barley and Brachypodium distachyon and characterised the wheat genes by heterologous expression and transcript analysis. Results The wheat, barley and Brachypodium genomes each contain orthologous copies of the GA20ox , GA3ox and GA2ox genes identified in rice, with the exception of OsGA3ox1 and OsGA2ox5 which are absent in these species. Some additional paralogs of 2-ODD genes were identified: notably, a novel gene in the wheat B genome related to GA3ox2 was shown to encode a GA 1-oxidase, named as TaGA1ox-B1. This enzyme is likely to be responsible for the abundant 1β-hydroxylated GAs present in developing wheat grains. We also identified a related gene in barley, located in a syntenic position to TaGA1ox-B1 , that encodes a GA 3,18-dihydroxylase which similarly accounts for the accumulation of unusual GAs in barley grains. Transcript analysis showed that some paralogs of the different classes of 2-ODD were expressed mainly in a single tissue or at specific developmental stages. In particular, TaGA20ox3 , TaGA1ox1 , TaGA3ox3 and TaGA2ox7 were predominantly expressed in developing grain. More detailed analysis of grain-specific gene expression showed that while the transcripts of biosynthetic genes were most abundant in the endosperm, genes encoding inactivation and signalling components were more highly expressed in the seed coat and pericarp. Conclusions The comprehensive expression and functional characterisation of the multigene families encoding the 2-ODD enzymes of the GA pathway in wheat and barley will provide the basis for a better understanding of GA-regulated development in these species. This analysis revealed the existence of a novel, endosperm-specific GA 1-oxidase in wheat and a related GA 3,18-dihydroxylase enzyme in barley that may play important roles during grain expansion and development.

The composition of proanthocyanidins in the testa (seed coat) of bread wheat was analyzed by thiolysis of PA oligomers from developing grain and found to consist of (+)‐catechin monomers, with a small amount of (+)‐gallocatechin. The average chain length of soluble PA stayed relatively constant between 10 and 20 days post‐anthesis, whereas that of unextractable PA increased over the same period, suggesting that increases in chain length might account for the insolubility of PAs from mature wheat grain. We carried out RNA‐Seq followed by differential expression analysis from dissected tissues of developing grain from red‐ and white‐grained near‐isogenic lines differing in the presence of an active R gene that encodes a MYB transcription factor involved in control of PA biosynthesis. In addition to genes already identified encoding chalcone synthase, chalcone isomerase, flavanone 3‐hydroxylase, and dihydroxyflavonoid 4‐reductase, we showed that wheat genes encoding phenylalanine ammonia lyase, flavonoid 3′,5′‐hydroxylase, leucoanthocyanidin reductase, and a glutathione S‐transferase (the orthologue of maize Bronze‐2) were more highly expressed in the red NIL. We also identified candidate orthologues of other catalytic and regulatory components of flavonoid biosynthesis in wheat.

Navan Fort is an iconic prehistoric Irish ceremonial centre and the legendary capital of Ulster. The fort has produced an exceptional pig-dominated faunal assemblage that also contained a barbary macaque skull. Dating from the 4 th to 1 st century BC, it is likely to be a ceremonial feasting centre that may have drawn people and their animals from across Ulster and beyond. This study uses a multi-isotope ( 87 Sr/ 86 Sr, δ 34 S, δ 13 C, δ 15 N) approach to identify non-local animals and reconstruct site catchment. New biosphere mapping means that isotope data can be more confidently interpreted and the combination of strontium and sulphur analysis has the potential to estimate origins. In the absence of human remains, fauna provide the best proxy for human movement. Results for the 35 analysed animals are wide-ranging, especially in terms of strontium (0.707–0.715), which has the largest range for an Irish site. Sulphur values are more restricted (13.1‰−17.1‰) but are high in the context of British and Irish data. Results provide clear evidence for animals (and thus people) coming from across Ulster and beyond, demonstrating the site’s wide catchment. Navan Fort was clearly a major ceremonial centre with far-reaching influence and hosted feasts that drew people and animals from afar.

Background Despite the importance of the Plasmodium berghei oocyst capsule protein (PbCap380) in parasite survival, very little is known about the orthologous Plasmodium falciparum capsule protein (PfCap380). The goal of this work was to study the growth of P. falciparum oocysts using PfCap380 as a developmental marker. Methods To study P. falciparum oocyst development using both in vivo (mosquito-derived) and in vitro (culture-derived) growth conditions, antibodies (polyclonal antisera) were raised against PfCap380. For studies on in vivo oocysts, mature P. falciparum gametocytes were fed to Anopheles stephensi mosquitoes. For studies on in vitro parasites, P. falciparum gametocytes were induced and matured for subsequent ookinete production. Ookinetes were purified and then tested for binding affinity to basal lamina components and transformation into early oocysts, which were grown on reconstituted basal lamia coated wells with novel oocyst media. To monitor in vivo oocyst development, immunofluorescence assays (IFA) were performed using anti-PfCap380 antisera on Pf -infected mosquito midguts. IFA were also performed on culture-derived oocysts to follow in vitro oocyst development. Results The anti-PfCap380 antisera allowed detection of early midgut oocysts starting at 2 days after gametocyte infection, while circumsporozoite protein was definitively observed on day 6. For in vitro culture, significant transformation of gametocytes to ookinetes (24%) and of ookinetes to early oocysts (85%) was observed. After screening several basal lamina components, collagen IV provided greatest binding of ookinetes and transformation into early oocysts. Finally, PfCap380 expression was observed on the surface of culture-derived oocysts but not on gametocytes or ookinetes. Conclusions This study presents developmental monitoring of P. falciparum oocysts produced in vivo and in vitro. The anti-PfCap380 antisera serves as an important reagent for developmental studies of oocysts from the mosquito midgut and also from oocyst culture using in vitro methodology. The present data demonstrate that PfCap380 is a useful marker to follow the development and maturation of in vivo and in vitro produced oocysts as early as 2 days after zygote formation. Further in vitro studies focused on oocyst and sporozoite maturation will support the manufacturing of whole sporozoites for malaria vaccines.