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Bison (Bison spp) are being reintroduced into semi-wild, spatially constrained herds across North America and Europe. Herd managers are concerned about gastrointestinal (GI) nematode parasites as they care for the health of their bison. We examine how demographics, grazing location, herd management, and anthelmintic treatments affect the fecal egg counts (FECs) of GI nematodes within a reintroduced Plains bison (Bison bison bison) herd in the Great Plains. Our results suggest that younger bison (<2 years of age) experience higher GI parasite eggs/oocysts per gram (epg/opg) and that some taxa are more prevalent throughout different periods of a bison's early years. Demographic findings suggest that calf and yearling (0–2 yrs age) bison have the highest FECs and that these decline until reaching a low in peak adulthood and thereafter (x > 6 yrs of age). FECs of both Trichuris spp. and particularly Nematodirus spp. were much more abundant, relatively, during the first year of a bison's life. This pattern was also true of Moniezia spp. and Eimeria spp., however, strongyle-type spp. FECs appeared to peak in relative abundance during the second year of life. Our data also indicate that FECs are influenced by differences in land-use histories of pastures previously grazed by cattle or by the proportion of frequent flooding in different pastures. Treatment results suggest that fenbendazole may more effective than moxidectin at lowering FECs of bison over the long-term, and lasting effects of at least one administered anthelmintic treatment. Multiplex PCR assays revealed that American bison share GI nematodes with cattle including: Ostertagia spp., Haemonchus placei, Cooperia onchophora , and Oesophagostomum spp, but did not detect the presence Trichostrongylus columbriformis. Our results may have wider conservation implications for reintroduction efforts of American bison, as well as the endangered European bison (Bison bonasus). [Display omitted] • Younger plains bison (<2 years of age) are more likely to have higher fecal egg counts of gastrointestinal nematodes. • Eimeria spp., Trichuris spp., Moniezia spp., and Nematodirus spp. are more prevalent in plains bison between zero and 1 years of age, while strongyle-type nematodes are more prevalent in bison between 1 and 2 years of age. • Both land-use histories and landscape features may influence higher fecal egg counts of gastrointestinal nematodes in plains bison. • Plains bison share many of the same type of gastrointestinal parasites found in cattle. • Fenbendazole anthelmintic was more effective than moxidectin treatments in reducing fecal egg counts of gastrointestinal parasites in plains bison. [ABSTRACT FROM AUTHOR]

Highlights: [•] Trichinella consists of a complex assemblage of no less than 12 different taxa. [•] Extant encapsulated Trichinella spp. appeared to within the last 20 million years. [•] The Trichinella genus diverged very early in the evolution of the nematode phylum. [•] Approximately 9,000 genes have been identified in the Trichinella genus. [•] Trichinella spp. developed different adaptive strategies. [ABSTRACT FROM AUTHOR]

Abstract: Incidences of H1N1 viral infections in Mainland China are collected by the Ministry of Health, the People''s Republic of China. The number of confirmed cases and the timing of these outbreaks from May 13 to July 22, 2009 were obtained and subjected to a novel mathematical model to simulate the infection profile (time vs number). The model was predicated upon the grey prediction theory which allows assignment of future trends using limited numbers of data points. During the period of our analysis, the number of confirmed H1N1 cases in Mainland China increased from 1 to 1772. The efficiency of our model to simulate these data points was evaluated using Sum of squares of error (SSE), Relative standard error (RSE), Mean absolute deviation (MAD) and Average relative error (ARE). Results from these analyses were compared to similar calculations based upon the grey prediction algorithm. Using our equation, defined herein as equation D–R, results showed that SSE=6742.00, RSE=10.69, MAD=7.07, ARE=2.47% were all consistent with the D–R algorithm performing well in the estimation of future trends of H1N1 cases in Mainland China. Calculations using the grey theory had no predictive value [ARE for GM(1,1)=−104.63%]. To validate this algorithm, we performed a second analysis using new data obtained from cases reported to the WHO and CDC in the US between April 26 and June 8, 2009. In like manner, the model was equally predictive. The success of the D–R mathematical model suggests that it may have broader application to other viral infections among the human population in China and may be modified for application to other regions of the world. [Copyright &y& Elsevier]

Abstract: Four species of Cryptosporidium are routinely found in cattle: Cryptosporidium parvum, Cryptosporidium bovis, Cryptosporidium ryanae, and Cryptosporidium andersoni. It is important to determine the species of Cryptosporidium in infected cattle because C. parvum is the only serious pathogen for humans as well as cattle. Identification of Cryptosporidium species and genotypes currently relies on molecular methods such as polymerase chain reaction (PCR) followed by restriction fragment length polymorphism (RFLP) or gene sequencing. Incorporation of these techniques in a routine veterinary diagnostic laboratory is cost prohibitive. As such, their applications are limited primarily to research and a few public health laboratories. To overcome this problem, a multiplex PCR assay was developed for simultaneously detecting the 4 species of Cryptosporidium that commonly infect cattle. This assay specifically identifies Cryptosporidium oocysts present in cattle feces, improves the detection of mixed infections, reduces the time and cost relative to current sequencing methods, and further demonstrates the shortcomings of sequencing as the definitive method for identification when analyzing samples containing mixed infections. [Copyright &y& Elsevier]

Abstract: In 1990, the Human Genome Sequencing Project was established. This laid the ground work for an explosion of sequence data that has since followed. As a result of this effort, the first complete genome of an animal, Caenorhabditis elegans was published in 1998. The sequence of Drosophila melanogaster was made available in March, 2000 and in the following year, working drafts of the human genome were generated with the completed sequence (92%) being released in 2003. Recent advancements and next-generation technologies have made sequencing common place and have infiltrated every aspect of biological research, including parasitology. To date, sequencing of 32 apicomplexa and 24 nematode genomes are either in progress or near completion, and over 600k nematode EST and 200k apicomplexa EST submissions fill the databases. However, the winds have shifted and efforts are now refocusing on how best to store, mine and apply these data to problem solving. Herein we tend not to summarize existing X-omics datasets or present new technological advances that promise future benefits. Rather, the information to follow condenses up-to-date-applications of existing technologies to problem solving as it relates to parasite research. Advancements in non-parasite systems are also presented with the proviso that applications to parasite research are in the making. [Copyright &y& Elsevier]

Abstract: Ostertagia ostertagi is a gastrointestinal parasitic nematode that affects cattle and leads to a loss of production. In this study, we present the first large-scale genomic survey of O. ostertagi by the analysis of expressed transcripts from three stages of the parasite: third-stage larvae, fourth-stage larvae and adult worms. Using an in silico approach, 2284 genes were identified from over 7000 expressed sequence tags and abundant transcripts were analyzed and characterized by their functional profile. Of the 2284 genes, 66% had similarity to other known or predicted genes while the rest were novel and potentially represent genes specific to the species and/or stages. Furthermore, a subset of the novel proteins were structurally annotated and assigned putative function based on orthologs in Caenorhabditis elegans and corresponding RNA interference phenotypes. Hence, over 70% of the genes were annotated using protein sequences, domains and pathway databases. Differentially expressed transcripts from the two larval stages and their functional profiles were also studied leading to a more detailed understanding of the parasite''s life-cycle. The identified transcripts are a valuable resource for genomic studies of O. ostertagi and can facilitate the design of control strategies and vaccine programs. [Copyright &y& Elsevier]

Abstract: A multiplex PCR assay for differentiating strongyle eggs from cattle has recently been described; however, the egg disruption and DNA extraction procedures, though effective, are inadequate for large studies or clinical application. The purpose of this research was to evaluate methods for disrupting trichostrongyle eggs, then assess commercial kits for extracting egg DNA using Ostertagia ostertagi as a model species. Egg disruption procedures tested included probe sonication, bath sonication, bead beating, boiling, microwaving, proteinase K/SDS digestion, freezing, and various combinations of the above with the incorporation of sodium dodecyl sulfate. These procedures were evaluated in conjunction with four commercial DNA extraction kits: DNA Stool mini kit and DNeasy Plant kit (Qiagen), Fast DNA kit (QBiogene), and the MAP extraction kit (Tetracore). Results showed that egg disruption was best accomplished with the bead beater and ceramic beads, resulting in 100% disruption within 1min. When DNA extraction was preceded by the isolation of eggs from feces, all procedures except the Fast DNA kit produced PCR-ready DNA from at least two eggs. The DNeasy Plant kit allowed consistent detection of DNA released from one egg. Due to the morphological similarities among trichostrongyle eggs in ruminants, strongyle eggs in equids, and hookworm eggs, the methods described herein may have broad application to other nematodes. [Copyright &y& Elsevier]

Abstract: There has been recent emphasis on developing better methods for detecting diseases of zoonotic and veterinary importance. This has been prompted by an increase in human disease agents detectable in environmental samples, the potential for bioterrorism, and the lowering of international trade barriers and expansion of personal travel, which are bringing previously considered exotic diseases to new geographical localities. To appreciate the complexities of developing detection methods and working with environmental samples, it is appropriate to review technologies currently in use, as well as those in development and presently limited to research laboratories. Discussion of parasite detection would not be possible without including methods for parasite sampling, concentration, and purification because it is often necessary to process large sample volumes prior to analysis, and no reliable methods are available for significantly amplifying parasites in vitro. Reviewing proven methods currently in use will provide a baseline for generating, accepting and implementing the more sensitive and specific methods under development today. [Copyright &y& Elsevier]

IL-4 and IL-13 are multi-functional cytokines with overlapping roles in the host defense against infection. Equally important in the regulation of IL-4 and IL-13 are their associated receptors. Though, their functional receptor complexes and signaling pathways are intricate and in some cases, share common elements, the specificity of the responses, nonetheless, resides in the structure and binding of the α-chain components. This report presents the cloning of the swine receptors IL-4Rα and IL-13Rα1 and the effects of parasite infection on their transcription. Pairwise alignment of predicted amino acid sequences indicates that the swine IL-13Rα1 is 86, 83, and 72% similar to canine, human and mouse sequences, respectively. Amino acid sequence conservation is appreciably lower between the swine IL-4Rα sequence and those from equine (72%), human (66%), and mouse (49%); however, noteworthy similarities were observed in their overall predicted secondary structures predominantly among the swine, equine, and human homologues. Relative levels of receptor mRNA in tissues from swine experimentally infected with the protozoan, Toxoplasma gondii (T. gondii) or the nematodes Ascaris suum (A. suum) or Trichuris suis (T. suis), which are known to induce Th1 or Th2 host responses, respectively, were measured by real-time PCR. Results indicated that within 14 days following infection, overall mRNA levels for IL-4Rα and IL-13Rα1 were elevated in T. gondii-infected animals and reduced in A. suum-infected animals. Levels of swIL-4Rα and swIL-13Rα1 mRNA in T. suis-infected animals varied coincidentally with the course of the infection and the location of the analyzed tissue. [Copyright &y& Elsevier]

Presently, little is known of the mechanism by which Trichinella penetrates and modulates reprogramming of muscle cells. In light of evidence demonstrating strong protective characteristics of antigens derived from this stage, understanding this process may shed light on potential targets for effective abatement of infection. To this end, a PCR-derived cDNA expression library was constructed using 0.5 μg of total RNA from Trichinella spiralis newborn larvae. The library consisted of >125,000 insert-containing clones. Approximately 40–50×103 clones were screened immunologically using sera from pigs experimentally infected with 7,000 Trichinella L1. Multiple clones reacting positively with the swine infection serum and encoding portions of a glutamic acid-rich protein were identified. Northern and Southern blots indicated at least two distinct genes that encoded the glutamic acid-rich proteins and that these genes were transcribed specifically in the newborn larvae stage. cDNA sequence data predicted open reading frames of 1,497 and 1,716 bp generating proteins of 498 amino acids and 571 amino acids, respectively. Both sequences consisted of approximately 39% glutamic acid and 16% serine residues, and differed by the presence of a 219 bp fragment present in the 1,716 bp sequence that was absent from the 1,497 bp sequence. PCR data indicated that additional isoforms exist within this gene family that are different in length from those described above. In addition, it was found that more than one isoform can exist within a single worm and that this pattern can vary between individual worms within a population. Mouse antibodies to recombinant antigen localised the glutamic acid-rich proteins to the periphery of the developing stichocyte cells within the newborn larvae consistent with the hypothesis that the newborn larval antigens are secreted. [Copyright &y& Elsevier]

Standard therapies including administration of potent antibiotics, aggressive fluid resuscitation and metabolic support have not been successful in relieving symptoms and reducing mortality associated with acute coliform mastitis. It is important to understand the pathophysiological response of the mammary gland to coliform infections when designing preventive or therapeutic regimens for controlling coliform mastitis. Our laboratory has previously shown that macrophages and polymorphonuclear neutrophils in milk express CD14 on their cell surface. In this study, we found that soluble CD14 (sCD14) is present in milk whey as a 46 kDa protein reacted with anti-ovine CD14 antibody. Additional functional studies found that: (1) under serum-free condition, complexes of LPS-recombinant bovine soluble CD14 (rbosCD14) induced activation of mammary ductal epithelial cells (as measured by changes in interleukin-8 (IL-8) mRNA level by competitive RT–PCR) at low concentrations of LPS after 6 or 24 h incubation (1–1000 ng/ml), whereas LPS alone did not induce activation of mammary ductal epithelial cells at the same concentrations, and (2) intramammary injection of low concentrations of LPS did not increase concentration of leukocytes in milk. In contrast, LPS–rbosCD14 complex containing the same concentration of LPS increased the concentration of leukocytes in the injected mammary gland at 12 and 24 h post-injection. These results indicate that rbosCD14 sensitizes mammary epithelial cells to low concentrations of LPS in vitro and in vivo. Endogenous sCD14 in milk may be important in initiating host responses to Gram-negative bacterial infections. [Copyright &y& Elsevier]

Available evidence suggests that Trichinella spiralis first originated in Asia and subsequently spread to the rest of the world. Notably limited genetic diversity in European T. spiralis isolates indicates that the parasite went through a dramatic genetic bottleneck at some point in its history. Did this genetic bottleneck result from the transport of a limited number of T. spiralis infected pigs from Asian centers of domestication, or was the parasite resident in Europe far earlier than the domestication of pigs there? In order to explore this hypothesis, we generated complete mitochondrial genomes and ribosomal DNAs from seventeen European T. spiralis isolates, six North American isolates and seven Asian isolates using next generation sequencing. A total of 13,858 base pairs of mitochondrial DNA and 7431 nucleotides of the nuclear ribosomal DNA sequence from each isolate were aligned and subjected to phylogenetic analysis using T. nelsoni as an outgroup. We confirmed that North American and European isolates were tightly clustered within a single "western clade" and all Chinese T. spiralis isolates were placed within a well-supported sister clade. These results indicate that European T. spiralis did not directly descend from extant Chinese parasite populations. Furthermore, the amount of nucleotide divergence between the two clades suggests that they diverged before pigs were domesticated. Over evolutionary time periods, Chinese and European T. spiralis were likely maintained as separate populations. The data presented here indicates the genetic bottleneck observed in European T. spiralis did not result from a small number of founders introduced with Chinese pigs in the recent past, but derives from an earlier bottleneck in host populations associated with the end of the last glacial maximum. • The intraspecific history of Trichinella spiralis was examined using phylogenetics. • Complete mitochondrial genomes and nuclear ribosomal RNA cassettes were sequenced. • Asian and European T. spiralis isolates were divided into sister clades. • European populations of T. spiralis have a long independent history from Asian populations. [ABSTRACT FROM AUTHOR]

Hybridization between two closely related but distinct genetic lineages may lead to homogenization of the two lineages with potentially novel phenotypes, or selective pressure to avoid hybridization if the two lineages are truly distinct. Trichinella nativa and Trichinella T6 are zoonotic nematode parasites which can be distinguished genetically despite occasional hybridization. Here, using an experimental murine model, we attempt to determine whether there are barriers to hybridization when sizeable numbers of each lineage are allowed to coinfect a host. Two mice were independently infected with equal numbers of T. nativa and T6. The offspring of these coinfections were genotyped at two microsatellite loci and one mitochondrial locus capable of distinguishing T. nativa from T6 genotypes. Among larvae in the F1 generation, offspring of every possible mating were encountered. Most larvae (63.6%) derived from T. nativa × T. nativa matings, while 21.1% of offspring were the product of T6 × T6 matings, and only 15.3% were hybrid offspring of T. nativa × T6 crosses, differing markedly from null expectations. In this experimental model, T. nativa and Trichinella T6 were able to mate, but ratios of offspring indicated pre- or post-zygotic barriers to hybridization that may include assortative mating, genetic incompatibilities, and/or differences in the fitness of offspring. These barriers would limit gene flow between these two lineages in a natural setting, serving as a barrier to their homogenization and promoting their persistence as distinct and separate entities. [ABSTRACT FROM AUTHOR]

Abstract: The genetic diversity within and among parasite populations provides clues to their evolutionary history. Here, we sought to determine whether mitochondrial and microsatellite DNA variation could be used to evaluate the extent of differentiation, gene flow and historical reproductive isolation among the freeze resistant parasites Trichinella nativa and the Trichinella T6 genotype infecting wolverines (Gulo gulo) in Nunavut, Canada. To this end, we genotyped Trichinella isolates derived from the diaphragms of 39 wolverines from this locale to reference strains of T. nativa and the Trichinella T6 genotype. Results showed that among a subset of 13 isolates examined, individuals resembled T. nativa in their mitochondrial DNA, but resembled the Trichinella T6 genotype when assayed at expansion segment V and the internal transcribed spacer of the nuclear rDNA. To adjudicate among these conflicting diagnoses, we further characterized each isolate at several nuclear microsatellite loci and again compared these to data from reference strains. Statistical assignment established that the nuclear genomes of most Nunavut isolates corresponded to those of the Trichinella T6 genotype; however, two isolates corresponded to T. nativa, and one isolate exhibited equal similarity to both reference strains. Taken as a whole, the evidence suggests that these isolates derive from the T. nativa matrilineage, but that their nuclear genomes resemble individuals previously designated as Trichinella T6. Assuming distinct lineages, this argues for cross-hybridization among these genotypes. Although introgression has occurred, recognizable genetic distinctions persist. One possibility is that selection disfavors the survival of hybrid offspring in most instances. Alternatively, the recent disappearance of glacial barriers might have increased contact, and therefore introgression. Broader geographic sampling will be required to determine the extent to which hybridization occurs beyond this particular geographic focus. [Copyright &y& Elsevier]

Abstract: Studying parasites of the genus Trichinella provides scientists of today many advantages. This is a group of zoonotic nematodes that circulates freely among wildlife hosts with one in particular, Trichinella spiralis that is exceptionally well adapted to domestic swine. Recent reports suggest that human infections from hunted animals are on the rise worldwide and numerous countries still experience problems with T. spiralis in their domestic food supplies. Trichinella is a genus whose members are easily propagated in the laboratories, have been used as models to investigate host–parasite relationships and parasitism among clade I organisms, and represent a poorly investigated link between the phylum Nematoda and other Metazoans. The importance of T. spiralis in better understanding the tree of life was so recognized that in 2004, its genome was carefully selected as one of only nine key non-mammalian organisms to be sequenced to completion. Since it was first discovered in 1835, this genus has expanded from being monospecific to eight species including four other genotypes of undetermined taxonomic rank. Inasmuch as discriminating morphological data have been scant, our understanding of the genus has been relegated to a compilation of molecular, biochemical and biological data. Herein, we provide a collection of information and up-to-date interpretations on the taxonomy, diagnostics, systematics, micro- and macroevolution, and the biogeographical and biohistorical reconstruction of the genus Trichinella. [Copyright &y& Elsevier]

Abstract: A non-isotopic single-strand conformation polymorphism (‘cold’ SSCP) technique has been assessed for the analysis of sequence variability in the expansion segment 5 (ES5) of domain IV and the D3 domain of nuclear ribosomal DNA within and/or among isolates and individual muscle (first-stage) larvae representing all currently recognized species/genotypes of Trichinella. Data are consistent with the ability of cold SSCP to identify intra-specific as well as inter-specific variability among Trichinella genotypes. The cold SSCP approach should be applicable to a range of other genetic markers for comparative studies of Trichinella populations globally. [Copyright &y& Elsevier]

• One third of the samples analyzed were positive for Clade V nematode DNA. • Ostertagia was found in 90 % of the sequenced samples. • Wild ruminants can act as reservoirs for maintaining infections in production herds. Gastrointestinal nematode (GIN) infections in cattle cause appetite suppression which leads to poor feed conversion, reduced weight gain and reduced milk production. Overuse and exclusive reliance on anthelmintic drugs has resulted in widespread resistance in many parasitic nematode species infecting livestock making control increasingly difficult. Wild ruminants are competent hosts of a number of nematode species that typically infect and are best adapted for cattle, sheep, and goats. Thus, the potential exists for wild ruminants to act as reservoirs in the translocation of domestic GIN, including those carrying anthelmintic resistance mutations as well as susceptible genotypes. The potential for parasite exchange is heightened by interfaces or ecotones between managed and wild rangelands, and by perturbations linked to climate warming that can increasingly alter the distributions of wild ungulates and their interactions with domestic and free-ranging ruminants. To investigate the extent to which wild ruminants harbour parasites capable of infecting domestic ruminants we first performed an epidemiological study of feces from wildlife hosts that spanned 16 states and included white-tailed deer (85 % of the samples), pronghorn, elk, mule deer, bighorn sheep, moose, cattle, and caribou across the United States. All samples were cultured to third stage larvae and nematode DNA was isolated and PCR amplified. Among the 548 wild ruminant samples received, 33 % (181 samples) were positive for nematode DNA, among which half (84 samples) contained DNA from GIN species commonly found in cattle. DNA from cattle GIN species was detected in 46 % of samples from the Northeast, 42 % from the Southeast, 10 % from the Midwest, 0 % from the Southwest and 11 % from the West. Deep amplicon sequencing of the ITS-2 rDNA indicated that Ostertagia and Trichostrongylus were present in 90 % and 69 % of the nematode DNA positive samples, respectively, whereas Haemonchus, Cooperia and Oesophagostomum were present in 26 %, 2 % and 10 % of the samples, respectively. These data clearly show that wild ruminants commonly harbour multiple parasite species whose primary hosts are domestic cattle, and suggest that further work is warranted to investigate their specific roles in the management of anthelmintic resistance. [ABSTRACT FROM AUTHOR]