Expression of genes related to ileal barrier function in heritage and modern broiler chickens.

1. An experiment was conducted to determine differences in the expression of genes encoding intestinal barrier proteins between fast, medium and slow-growing chickens. Chicken breeds Athens Canadian Random Bred (ACRB), Longenecker's Heritage (LHR), RedBro, Hubbard H1 (HH1), Cobb500 and Ross708 were raised from hatch for 35 d.2. Ileal samples were collected at embryonic day E19 (-2 days post-hatch), hatch and d 7, 14, 21, 28 and 35 post-hatch to assess the expression of genes encoding tight junction proteins (claudins, CLDN; occludin, OCLN; zonula occludens, ZO; and junctional adhesion molecules, JAM), mucin (Muc2), immunoglobulin A (IgA), polymeric immunoglobulin receptor (pIgR) and fatty acid binding protein (FABP2).3. Expression of CLDN-1 was increased (p < 0.0001) in LHR compared to Cobb500 while CLDN-5 was increased (p < 0.0001) in ACRB, HH1, RedBro and Ross708 compared to LHR as well as in ACRB compared to Cob500. Occludin was upregulated (p = 0.01) in ACRB and LHR compared to Ross708 at d 14 post-hatch. Expression of ZO-1 was upregulated (p = 0.001) in LHR compared to Ross708, HH1 and Cobb500. Tight junction genes, except CLDN-4, JAM-2 and JAM-3 were downregulated (p < 0.0001) at hatch and d 7 post-hatch. Expression of Muc2 was increased (p < 0.0001) in LHR compared to RedBro and from -2 d to d 7 post-hatch.4. Immunoglobulin A was increased (p = 0.001) in LHR compared to Ross708 and HH1 at -2 d post-hatch and in LHR compared to ACRB, Cobb500 and Ross708 at hatch. In addition, IgA expression was increased in all breeds at d 14 post-hatch while pIgR was upregulated (p = 0.02) in Cobb500 and Ross708 compared to ACRB, HH1, LHR and RedBro at hatch.5. The gene expression patterns suggest that selection for growth may have not induced changes in junctional complexes and immune defence genes. However, the results confirmed that the expression of these genes is age dependent.

Expression of genes associated with nutrient uptake in intestines of chickens with different growth potentials show temporal changes but are not correlated with growth.

1. The study was designed to compare the expression of genes that encode proteins located at either the brush border (BB) or basolateral (BL) of the gut epithelium among fast and slow-growing broilers.2. Six broiler breeds with different growth capacities were used: Ross 708, Hubbard H1 (HH1), Cobb 500, Longenecker's Heritage (LHR), Red-Bro, and the Athens Canadian Randombred Control (ACRB). Birds were sampled between embryonic day (ED) 19 and day 35 post-hatch (PH).3. Performance parameters indicated that Ross 708, HH1, and Cobb 500 had the highest body weights (BW) while ACRBs had the lowest.4. Quantitative RT-PCR was performed on 13 genes encoding proteins associated with nutrient processing and uptake. Statistical analysis was carried out (ANOVA) for eight BB genes: Aminopeptidase N (APN), four amino acid transporters, (ATBo,+, BoAT, bo,+AT, EAAT3) a di- and tri-peptide transporter (PepT1), and two sugar transporters (GLUT5 and SGLT1). Analysis of four amino acid transporters (CAT1, CAT2, LAT1, and γ+LAT1), and a single sugar transporter (GLUT2) associated with BL was carried out.5. Four BB associated genes (APN, EAAT3, BoAT, and b0,+AT) in the small intestine were negatively correlated with growth.6. In most cases, genes encoding BB proteins increased in expression over time (P < 0.05) in the small intestine, while, in the caeca, the expression decreased (P < 0.05). The mRNA of BL-associated proteins showed decreased (P < 0.05) expression over time in all gut segments, with exception of GLUT2, which increased in expression in the small intestine.7. The temporal changes in gene expression were consistent among bird lines and BB associated genes tended to increase over time, while BL associated genes tended to decrease over time. Correlation analysis indicated that mRNA expression of nutrient transporter genes may not be a good predictor of growth potential.

The effect of delayed feeding post-hatch on caeca development in broiler chickens.

1. Broiler chicks are frequently deprived of food up to 72 h due to uneven hatching rates, management procedures and transportation to farms. Little is known about the effect of delayed feeding due to extended hatching times on the early neonatal development of the caeca. Therefore, the objective of this study was to investigate the developmental changes and effects of a 48-h delay in feed access immediately post-hatch (PH) on the caeca.2. After hatch, birds (Ross 708) were randomly divided into two treatment groups (n = 6 battery pen/treatment). One group (early fed; EF) received feed and water immediately after hatch, while the second group (late fed; LF) had access to water but had delayed access to feed for 48 h. Contents averaging across all regions of the caeca were collected for mRNA expression as well as for histological analysis at -48, 0, 4 h PH and then at 1, 2, 3, 4, 6, 8, 10, 12 and 14 days PH.3. Expression of MCT-1 (a nutrient transporter), Cox7A2 (related to mitochondrial function) IgA, pIgR, and ChIL-8 (immune function) genes was affected by delayed access to feed that was dependent by the time PH. Expression of immune and gut barrier function-related genes (LEAP2 and MUC2, respectively) was increased in LF group. There was no effect of feed delay on expression of genes related to mitochondrial functions in the caeca, although developmental changes were observed (ATP5F1B, Cox4|1). Caecal mucus and muscle thickness were affected by delayed access to feed during caeca development.4. The data suggested a limited effect of delayed feed access PH on the developmental changes in caecal functions. However, the caeca seemed to be relatively resistant to delayed access to feed early PH, with only a few genes affected.

Differential expression of intestinal nutrient transporters and host defense peptides in Eimeria maxima-infected Fayoumi and Ross chickens.

Fayoumi chickens are believed to be more disease resistant compared to commercial broiler chickens. The objective of this study was to compare mRNA expression of intestinal nutrient transporters, digestive enzymes, and host defense peptides (HDP) between Eimeria maxima-challenged Fayoumi and Ross broiler chickens. At 21 d of age, Ross broilers and Fayoumi lines M5.1 and M15.2 were challenged with 1,000 E. maxima oocysts. Control birds were not challenged. Duodenum, jejunum, and ileum were sampled (n = 6) at 7 d post challenge. Gene expression was analyzed using relative quantification PCR. Data were analyzed by ANOVA and significance level was set at P < 0.05. There was numerical, but not statistically significant, differential weight gain depression for Ross (15%) and Fayoumi lines M5.1 (21%) and M15.2 (22%) and significant line-specific changes in gene expression. For nutrient transporters, there was downregulation of mRNA for the brush border membrane, amino acid transporters b0,+AT/rBAT, BoAT, and EAAT3 in different segments of the small intestine of Ross and both lines of Fayoumi chickens, indicating that E. maxima challenge likely caused a decrease in nutrient uptake. For HDP, there was downregulation of avian beta defensin (AvBD) 1, 6, 10, 12, and 13 mRNA in the jejunum of the 2 Fayoumi lines, but no change in the Ross broilers. In the duodenum, there was upregulation of AvBD10 mRNA in the Ross and both Fayoumi lines and additionally upregulation of AvBD11, 12, and 13 mRNA in only Fayoumi line M15.2. Liver expressed antimicrobial peptide 2 (LEAP2) mRNA was downregulated in the duodenum and jejunum of Ross and Fayoumi line M5.1 but not in Fayoumi line M15.2. The homeostatic, non-challenged levels of AvBD mRNA were greater in Fayoumi line M15.2 than Ross and Fayoumi line M5.1 in the duodenum and ileum. This study demonstrates tissue- and genetic line-specific transcriptional responses to E. maxima, highlights novel potential candidate genes for response to coccidiosis, and confirms a role for several previously reported genes in response to coccidiosis.

Expression of host defense peptides in the intestine of Eimeria-challenged chickens.

Avian coccidiosis is caused by the intracellular protozoan Eimeria, which produces intestinal lesions leading to weight gain depression. Current control methods include vaccination and anticoccidial drugs. An alternative approach involves modulating the immune system. The objective of this study was to profile the expression of host defense peptides such as avian beta-defensins (AvBDs) and liver expressed antimicrobial peptide 2 (LEAP2), which are part of the innate immune system. The mRNA expression of AvBD family members 1, 6, 8, 10, 11, 12, and 13 and LEAP2 was examined in chickens challenged with either E. acervulina, E. maxima, or E. tenella. The duodenum, jejunum, ileum, and ceca were collected 7 d post challenge. In study 1, E. acervulina challenge resulted in down-regulation of AvBD1, AvBD6, AvBD10, AvBD11, AvBD12, and AvBD13 in the duodenum. E. maxima challenge caused down-regulation of AvBD6, AvBD10, and AvBD11 in the duodenum, down-regulation of AvBD10 in the jejunum, but up-regulation of AvBD8 and AvBD13 in the ceca. E. tenella challenge showed no change in AvBD expression in any tissue. In study 2, which involved challenge with only E. maxima, there was down-regulation of AvBD1 in the ileum, AvBD11 in the jejunum and ileum, and LEAP2 in all 3 segments of the small intestine. The expression of LEAP2 was further examined by in situ hybridization in the jejunum of chickens from study 2. LEAP2 mRNA was expressed similarly in the enterocytes lining the villi, but not in the crypts of control and Eimeria challenged chickens. The lengths of the villi in the Eimeria challenged chickens were less than those in the control chickens, which may in part account for the observed down-regulation of LEAP2 mRNA quantified by PCR. Overall, the AvBD response to Eimeria challenge was not consistent; whereas LEAP2 was consistently down-regulated, which suggests that LEAP2 plays an important role in modulating an Eimeria infection.

The mRNA expression of amino acid and sugar transporters, aminopeptidase, as well as the di- and tri-peptide transporter PepT1 in the intestines of Eimeria infected broiler chickens.

Coccidiosis in chickens is caused by infection of gut epithelial cells with protozoan parasites of the genus Eimeria This disease causes losses to the poultry industry since infected birds fail to gain weight as rapidly as non-infected birds and efficiency of feed conversion is compromised. For the present study the effect of Eimeria on expression of components of amino acid and sugar uptake mechanisms was determined. Broiler chicks were infected with Eimeria maxima, which infects the jejunum; Eimeria acervulina, which infects the duodenum; or Eimeria tenella, which infects the ceca. Sections of the jejunum, duodenum, and ceca (depending on species of Eimeria) were taken at several time points between d zero and 14 post infection (PI) for mRNA expression analysis. Genes examined included one digestive enzyme, 7 peptide and amino acid transporters located on the brush border, 8 transporters located at the basolateral surface of the gut epithelium, and 5 sugar transporters. All 3 Eimeria species examined caused decrease in expression of brush border transporters particularly at d 5 to 7 PI, which corresponds to the time when pathology is greatest. The same pattern was seen in expression of sugar transporters. However, the expression of basolateral transporters differed among species. Eimeria tenella infection resulted in decreased expression of all basolateral transporters, while E. maxima infection caused increased expression of 2 genes and slight decrease in expression of the remaining 5 genes. Infection with E. acervulina resulted in increased expression at the height of infection of all but one basolateral transporter. In conclusion, Eimeria infection causes a general decrease in gene expression of sugar transporter and brush border AATs at the height of infection. However the expression of basolateral transporters is increased in E. maxima and E. acervulina infected birds. It is possible that decreased expression of brush border transporters in combination with increased expression of basolateral transporters leads to decrease of nutrients available for the parasite, thus limiting parasite reproduction.

Impact of broiler egg storage on the relative expression of selected blastoderm genes associated with apoptosis, oxidative stress, and fatty acid metabolism.

Cool temperature storage of eggs prior to incubation is a frequent practice by commercial broiler hatcheries. However, continued storage beyond 7 d leads to a progressive increase in the rate of early embryonic mortality. In this study, we examined the relative expression of 31 genes associated with fatty acid metabolism (8), apoptosis (7), and oxidative stress (16) pathways to better understand the basis of embryo mortality during egg storage. A total of 642 broiler eggs in 2 separate trials were subjected to the following egg treatments: stored 4 d (Control 1, C1); stored 21 d but subjected to short periods of incubation during egg storage (SPIDES); stored un-manipulated 21 d (NonSPIDES, NS); and stored 4 d then incubated for 10 h to advance the embryos to the same developmental stages as the SPIDES embryos (Control 2, C2). Hatchability trials (277 eggs) confirmed the efficacy of SPIDES compared to NS treatments in both trials. To determine relative expression of 31 selected genes, 365 blastoderms were isolated, staged, and flash frozen in batches of 5 to 10 blastoderms per vial (7 vials per egg treatment) prior to RNA extractions. Analysis of gene expression was performed using qRT-PCR and the results presented as relative expression normalized to C1. The relative expression of genes in which the SPIDES and C2 treatments were significantly up- or down-regulated in tandem indicated that the stage-specific expression of those genes was maintained by the SPIDES treatment. This study provides the relative gene expressions of blastodermal cells before and after prolonged egg storage as well as insight as to how SPIDES impacts blastodermal cell gene expression.

Changes in expression of an antimicrobial peptide, digestive enzymes, and nutrient transporters in the intestine of E. praecox-infected chickens.

Coccidiosis is a major intestinal disease of poultry, caused by several species of the protozoan Eimeria. The objective of this study was to examine changes in expression of digestive enzymes, nutrient transporters, and an antimicrobial peptide following an Eimeria praecox challenge of chickens at days 3 and 6 post-infection. Gene expression was determined by real-time PCR and analyzed by one-way ANOVA. In the duodenum, the primary site of E. praecox infection, a number of genes were downregulated at both d3 and d6 post-infection. These genes included liver expressed antimicrobial peptide 2 (LEAP2), the cationic (CAT1), anionic (EAAT3), and L-type (LAT1) amino acid transporters, the peptide transporter PepT1 and the zinc transporter ZnT1. Other transporters were downregulated either at d3 or d6. At both d3 and d6, there was downregulation of B(o)AT and CAT1 in the jejunum and downregulation of LEAP2 and LAT1 in the ileum. LEAP2, EAAT3, and ZnT1 have been found to be downregulated following challenge with other Eimeria species, suggesting a common cellular response to Eimeria.

Characterization of the Eimeria maxima sporozoite surface protein IMP1.

The purpose of this study was to characterize Eimeria maxima immune-mapped protein 1 (IMP1) that is hypothesized to play a role in eliciting protective immunity against E. maxima infection in chickens. RT-PCR analysis of RNA from unsporulated and sporulating E. maxima oocysts revealed highest transcription levels at 6-12h of sporulation with a considerable downregulation thereafter. Alignment of IMP1 coding sequence from Houghton, Weybridge, and APU-1 strains of E. maxima revealed single nucleotide polymorphisms that in some instances led to amino acid changes in the encoded protein sequence. The E. maxima (APU-1) IMP1 cDNA sequence was cloned and expressed in 2 different polyHis Escherichia coli expression vectors. Regardless of expression vector, recombinant E. maxima IMP1 (rEmaxIMP1) was fairly unstable in non-denaturing buffer, which is consistent with stability analysis of the primary amino acid sequence. Antisera specific for rEmaxIMP1 identified a single 72 kDa protein or a 61 kDa protein by non-reducing or reducing SDS-PAGE/immunoblotting. Immunofluorescence staining with anti-rEmaxIMP1, revealed intense surface staining of E. maxima sporozoites, with negligible staining of merozoite stages. Immuno-histochemical staining of E. maxima-infected chicken intestinal tissue revealed staining of E. maxima developmental stages in the lamnia propia and crypts at both 24 and 48 h post-infection, and negligible staining thereafter. The expression of IMP1 during early stages of in vivo development and its location on the sporozoite surface may explain in part the immunoprotective effect of this protein against E. maxima infection.

Expression of digestive enzymes and nutrient transporters in Eimeria-challenged broilers.

Avian coccidiosis is a disease caused by the intestinal protozoa Eimeria. The site of invasion and lesions in the intestine is species-specific, for example E. acervulina affects the duodenum, E. maxima the jejunum, and E. tenella the ceca. Lesions in the intestinal mucosa cause reduced feed efficiency and body weight gain. The growth reduction may be due to changes in expression of digestive enzymes and nutrient transporters in the intestine. The objective of this study was to compare the expression of digestive enzymes, nutrient transporters and an antimicrobial peptide in broilers challenged with either E. acervulina, E. maxima or E. tenella. The genes examined included digestive enzymes (APN and SI), peptide and amino acid transporters (PepT1, ASCT1, b(0,+)AT/rBAT, B(0)AT, CAT1, CAT2, EAAT3, LAT1, y(+)LAT1 and y(+)LAT2), sugar transporters (GLUT1, GLUT2, GLUT5 and SGLT1), zinc transporter (ZnT1) and an antimicrobial peptide (LEAP2). Duodenum, jejunum, ileum and ceca were collected 7 days post challenge. E. acervulina challenge resulted in downregulation of various nutrient transporters or LEAP2 in the duodenum and ceca, but not the jejunum or ileum. E. maxima challenge produced both downregulation and upregulation of nutrient transporters and LEAP2 in all three segments of the small intestine and ceca. E. tenella challenge resulted in the downregulation and upregulation of nutrient transporters and LEAP2 in the jejunum, ileum and ceca, but not the duodenum. At the respective target tissue, E. acervulina, E. maxima and E. tenella infection caused common downregulation of APN, b(0,+)AT, rBAT, EAAT3, SI, GLUT2, GLUT5, ZnT1 and LEAP2. The downregulation of nutrient transporters would result in a decrease in the efficiency of protein and polysaccharide digestion and uptake, which may partially explain the weight loss. The downregulation of nutrient transporters may also be a cellular response to reduced expression of the host defense protein LEAP2, which would diminish intracellular pools of nutrients and inhibit pathogen replication.

Expression of digestive enzymes and nutrient transporters in Eimeria acervulina-challenged layers and broilers.

Avian coccidiosis is a disease caused by intestinal protozoa in the genus Eimeria. Clinical signs of coccidiosis include intestinal lesions and reduced feed efficiency and BW gain. This growth reduction may be due to changes in expression of digestive enzymes and nutrient transporters in the intestine. The objective of this study was to examine the differential expression of digestive enzymes, transporters of amino acids, peptides, sugars, and minerals, and an antimicrobial peptide in the small intestine of Eimeria acervulina-infected broilers and layers. Uninfected broilers and layers, in general, expressed these genes at comparable levels. Some differences included 3-fold and 2-fold greater expression of the peptide transporter PepT1 and the antimicrobial peptide LEAP2 (liver expressed antimicrobial peptide 2), respectively, in the jejunum of layers compared with broilers and 17-fold greater expression of LEAP2 in the duodenum of broilers compared with layers. In the duodenum of Eimeria-infected broilers and layers, there was downregulation of aminopeptidase N; sucrase-isomaltase; the neutral, cationic, and anionic amino acid transporters b(o,+)AT/rBAT, B(o)AT, CAT2, and EAAT3; the sugar transporter GLUT2; the zinc transporter ZnT1; and LEAP2. In the jejunum of infected layers there was downregulation of many of the same genes as in the duodenum plus downregulation of PepT1, b(o,+)AT/rBAT, and the y(+) L system amino acid transporters y(+) LAT1 and y(+) LAT2. In the ileum of infected layers there was downregulation of CAT2, y(+)LAT1, the L type amino acid transporter LAT1, and the sugar transporter GLUT1, and upregulation of APN, PepT1, the sodium glucose transporter SGLT4, and LEAP2. In E. acervulina-infected broilers, there were no gene expression changes in the jejunum and ileum. These changes in intestinal digestive enzyme and nutrient transporter gene expression may result in a decrease in the efficiency of protein digestion, uptake of important amino acids and sugars, and disruption of mineral balance that may affect intestinal cell metabolism and Eimeria replication.

Analysis of giardin expression during encystation of Giardia lamblia.

The present study analyzed giardin transcription in trophozoites and cysts during encystation of Giardia lamblia . Encystment was induced using standard methods, and the numbers of trophozoites and cysts were counted at various time points during the process. At all time points, RNA from both stages were assayed for levels of alpha2-, beta-, and delta-giardin mRNA as well as for cyst wall protein 3 (CWP3) mRNA using quantitative RT-PCR. In encystation medium, the number of G. lamblia trophozoites decreased, while the number of cysts increased between 0 and 72 hr. In trophozoites, alpha2- and beta-giardin transcription decreased over time, while delta-giardin transcription remained unchanged during the same time period. CWP3 transcription exhibited a slight increase in trophozoites at 8 hr, followed by a decrease at subsequent time points. Expression of alpha2-giardin increased at 48 hr in cysts followed by decreased expression at 72 hr, while beta- and delta-giardin expression was unchanged during encystation. CWP3 transcription gradually decreased from 24-72 hr in cysts. Consistent with previous studies, giardin proteins appeared to be disassembled into amorphous structures inside cysts during encystation. These findings represent the first analysis of giardin transcription in separate populations of trophozoites and cysts during encystation and indicate differential regulation of giardin mRNA expression by these developmental stages.

Gray wolf (Canis lupus) is a natural definitive host for Neospora caninum.

The gray wolf (Canis lupus) was found to be a new natural definitive host for Neospora caninum. Neospora-like oocysts were found microscopically in the feces of three of 73 wolves from Minnesota examined at necropsy. N. caninum-specific DNA was amplified from the oocysts of all three wolves. Oocysts from one wolf were infective for the gamma interferon gene knock out (KO) mice. Viable N. caninum (designated NcWolfUS1) was isolated in cell cultures seeded with tissue homogenate from the infected mouse. Typical thick walled tissue cysts were found in outbred mice inoculated with the parasite from the KO mouse. Tissue stages in mice stained positively with N. caninum-specific polyclonal antibodies. Our observation suggests that wolves may be an important link in the sylvatic cycle of N. caninum.

Molecular characterization and phylogenetic analysis of Eimeria from turkeys and gamebirds: implications for evolutionary relationships in Galliform birds.

In order to determine the evolutionary relationships among Eimeria species that parasitize birds of the Galliformes, the 18s rDNA gene and a portion of the cytochrome oxidase subunit 1 (cox-1) were amplified from Eimeria species isolated from turkeys, chukars, and pheasants. The phylogenetic analysis of these sequences suggests that species infecting chickens are polyphyletic and, therefore, do not all share a direct common ancestor. Both the 18s rDNA and the cox-1 sequences indicate that Eimeria tenella and Eimeria necatrix are more closely related to Eimeria of turkeys and pheasants than to other species that infect the chicken. It is, therefore, likely that the chicken Eimeria spp. represent 2 separate ancestral colonizations of the gut, one of which comprises E. tenella and E. necatrix that infect the ceca, while the other includes Eimeria acervulina, Eimeria brunetti, Eimeria maxima, and Eimeria mitis, which infect the upper regions of the intestine.

Comparison of Eimeria species distribution and salinomycin resistance in commercial broiler operations utilizing different coccidiosis control strategies.

The purpose of the present study was to evaluate the species composition and salinomycin sensitivity of Eimeria oocysts isolated from commercial broiler farms that differed by means of coccidiosis control (anticoccidial drugs [ACD] vs. live oocyst vaccines [VAC]). A comparison of Eimeria species composition and salinomycin sensitivity was also made before and after a producer switched from salinomycin to live oocyst vaccines. In general, no significant difference was observed in the concentration of Eimeria spp. oocysts in litter from VAC-utilizing farms compared to litter from ACD-utilizing farms. Application of PCR-based methods to detect coccidia found that Eimeria species distribution in litter from VAC operations more closely resembled the species composition in the live oocyst vaccines. Drug sensitivity testing found that Eimeria oocysts from VAC operations displayed greater salinomycin sensitivity as measured by weight gain and feed conversion efficiency compared to oocysts from ACD farms. These findings provide additional evidence for the usefulness of live oocyst vaccines to restore ionophore sensitivity in poultry operations that contain an ionophore-resistant population of Eimeria spp. oocysts.

Metam sodium reduces viability and infectivity of Eimeria oocysts.

Metam sodium (MS, sodium N-methyldithiocarbamate) is a widely used soil pesticide. Fumigation or chemical sterilization of poultry litter containing infectious oocysts could be an effective strategy to block the transmission of avian coccidia. In the current study, the effect of MS on the viability and infectivity of ocysts was investigated. The development of isolated, unsporulated oocysts of both Eimeria tenella and Eimeria maxima was inhibited, in a dose-related manner (IC(50) 8 to 14 microg/ml), by exposure to aqueous MS. Most treated oocysts failed to develop beyond early stages of sporulation. To determine the effect of MS on infectivity, isolated oocysts of E. tenella , Eimeria acervulina , and E. maxima were exposed for 24 hr to aqueous concentrations of MS ranging from 0 to 1,000 microg/ml. Treated oocysts were inoculated into chickens, and parameters of coccidiosis infection were compared to chickens inoculated with equal numbers of untreated oocysts. In a dose-related manner, MS significantly reduced the infectivity of oocysts with maximum effect observed at a dose of 300 microg/ml. When a mixture of oocysts containing 3 coccidian species was exposed to 300 microg/ml MS, from 0 to 24 hr, infectivity of oocysts was significantly reduced after a minimum of 12 hr of exposure. Treatment of aqueous slurries of litter samples obtained from commercial poultry houses, with 300 microg/ml MS for 24 hr, prevented the sporulation of eimerian oocysts in the litter samples relative to untreated control samples. The results indicate that MS could be used to reduce coccidial contamination of poultry litter.

Detection and comparison of Giardia virus (GLV) from different assemblages of Giardia duodenalis.

Five assemblages of Giardia duodenalis were identified from cysts in cattle, dog, cat, sheep, and reindeer feces using ribosomal DNA (rDNA) sequencing. Assemblage A was present in cattle and reindeer feces, Assemblages C and D were present in dog feces, Assemblage E was present in cattle and sheep feces, and Assemblage F was present in cat feces. Giardia virus, originally referred to as Giardia lamblia virus (GLV), is a double-stranded RNA virus. Primers designed for the GLV capsid protein gene identified GLV sequences in G. lamblia from a reindeer (Assemblage A) and from a dog (Assemblage C). Two distinct GLV sequences were identified in the dog specimen and 1 sequence was identified in the reindeer specimen. None of these GLV sequences was identical with previously published GLV sequences. It appears that GLVs are genetically diverse and that more than 1 virion can be present in a single sample. Because many of the specimens that contained cysts were found to be negative for GLV, it appears that this test for capsid protein is of limited value for the purposes of detecting G. lamblia.

Co-infection of chickens with Eimeria praecox and Eimeria maxima does not prevent development of immunity to Eimeria maxima.

Previous studies revealed an ameliorating effect of Eimeria praecox on concurrent E. maxima infection, such that weight gain, feed conversion ratio, and intestinal lesions were nearly identical to uninfected or E. praecox-infected controls. The purpose of the present study was to determine if protective immunity against E. maxima challenge infection developed in chickens infected with both E. praecox and E. maxima. Day-old chickens were infected with 10(3)E. praecox, 10(3)E. maxima, or a mixture of 10(3)E. praecox and 10(3)E. maxima oocysts. Chickens were then challenged at 4 weeks of age with 5x10(4)E. praecox or 5x10(3)E. maxima oocysts and clinical signs of coccidiosis were assessed 7 days post-challenge. Relative to non-challenged controls, naïve chickens or chickens immunized with E. praecox displayed a 32-34% weight gain depression after challenge with 5x10(3)E. maxima oocysts. In contrast, chickens immunized with either E. maxima oocysts alone or a combination of E. praecox and E. maxima oocysts displayed complete protection against lower weight gain associated with E. maxima challenge. Also, protection against decreased feed conversion ratio and intestinal lesions was observed in single E. maxima- or dual E. maxima+E. praecox-immunized chickens. These findings indicate that co-infection of chickens with E. maxima and E. praecox does not prevent development of immunity against E. maxima or E. praecox challenge.

Eimeria praecox infection ameliorates effects of Eimeria maxima infection in chickens.

The effect of Eimeria praecox on concurrent Eimeria maxima infection was studied in susceptible chickens. Clinical signs of coccidiosis were assessed in single E. praecox or E. maxima infections and compared to dual infection with both Eimeria species. Groups infected solely with 10(4)E. maxima oocysts displayed weight gains that were 48% of weight gain in uninfected controls. Weight gain in chickens infected only with 10(4)E. praecox oocysts was 90% of uninfected controls. Average weight gain in chickens infected with both E. maxima and E. praecox was 79% of controls, and showed no significant difference (P>0.05) from weight gain in E. praecox-infected chickens. Feed utilization (feed conversion ratio, FCR) in chickens infected with both species showed no significant difference (P>0.05) from FCR in non-infected controls or chickens infected with E. praecox alone; all showing a significant difference (P<0.05) from FCR in chickens infected solely with E. maxima. Although E. praecox did not appear to have a negative effect on weight gain and FCR, it did cause a significant decrease in serum carotenoids. Analysis of oocysts excreted by chickens during dual infection showed little effect of E. praecox on E. maxima oocyst production.