Sensitivity of field isolates of Eimeria acervulina and E. maxima from three regions in Brazil to eight anticoccidial drugs.

Rotation with different active ingredients is among the most effective and recommended strategies to preserve the efficacy of anticoccidial drugs and reduce the emergence of resistance. Tools such as anticoccidial sensitivity tests (ASTs) are ideally used to make rational rotation programs and bring benefits to production. The objective of this study was to evaluate the sensitivity of E. acervulina (EA) and E. maxima (EM) from 3 different regions in Brazil, by using four ASTs. Feces samples weighing 6 to 7 kg were collected in the regions of São Paulo, Paraná, and Minas Gerais. Prevalent oocysts from feces were filtered, identified, and quantified to conduct 2 ASTs with EA and 2 with EM. The same experimental design was used in every AST (4 replicates per treatment, with 6 birds each, for a total of 240 birds). Treatment groups were a nonchallenged and nonmedicated control group (T1), a challenged and nonmedicated control group (T2), and the other groups challenged and treated with the following compounds: lasalocid (90 ppm - T3), maduramycin (6 ppm - T4), decoquinate (30 ppm - T5), nicarbazin+semduramicin (66 ppm - T6), monensin (110 ppm - T7), salinomycin (66 ppm - T8), narasin+nicarbazin (100 ppm - T9), and nicarbazin (125 ppm - T10). At the end of each AST (20 d), the percent change (delta value) between the treated group (T3 to T10) and the control group (T2) was calculated for the following variables: body weight gain, feed conversion ratio, lesion score, and an indicator of percentage of optimal anticoccidial activity (POAA) that included T2. Different sensitivity levels of EA and EM isolates could be identified. As a whole, drugs from T5 and T3 groups showed higher delta values when compared to other compounds, whereas the lowest sensitivity levels of these isolates were observed in groups T4 and T7. Despite some limiting factors, ASTs can be a good tool for strategic selection of anticoccidial drugs in order to maintain efficacy and extend the lifespan of these molecules.

Microencapsulated and uncoated butyric acid as alternative additives to the regeneration of intestinal mucosa in broilers challenged with Eimeria spp.

1. The effect of microencapsulated and uncoated butyric acid as an alternative to antibiotics on performance, intestinal morphology and regeneration of intestinal mucosa was studied in birds experimentally infected with Eimeria spp. 1 to 42 d-old.2. A total of 1,320 male Cobb® broiler chicks were allocated to one of five treatments in a completely randomised design, comprising a negative control, uncoated butyric acid (UA), microencapsulated butyric acid (MA), combined U + M butyric acid and a positive control (antibiotic+anticoccidial) in six replications. At 16 d-old, the birds were inoculated orally with 0.5 ml of a solution containing an Eimeria spp. pool.3. At 21 d of age, the birds receiving butyric acid alone had higher body weight gain (BWG) and feed intake (FI) compared to those supplemented with the blend of acids. For the total rearing period, in all variables, the positive control performed best (P < 0.001).4. At 14 d of age, birds that received diets containing UA had a deeper crypt depth in the jejunum than those fed diets containing microencapsulated acid (P = 0.0194). At 21 d of age, the birds fed the acids had higher villi (P = 0.0058) in the duodenum, compared to the negative control group.5. Supplementation with microencapsulated acid contributed to the intestinal health and recovery of post-challenge birds, but did not result in improvements in performance.

Agaricus subrufescens and Pleurotus ostreatus mushrooms as alternative additives to antibiotics in diets for broilers challenged with Eimeria spp.

1. The effect of A. subrufescens and P. ostreatus mushrooms as an alternative to antibiotics (avilamycin or monensin sodium) on performance, intestinal morphometry, immunity, and biochemical profile of broilers challenged with Eimeria spp. was studied from 1 to 42 d old. A total of 900 male Cobb® broiler chicks were distributed, according to a completely randomised design, into five treatments with six replicates each.2. The treatments consisted of: negative control (NC) - basal diet (BD) with no anticoccidial or antibiotic (non-challenged birds); negative control challenged (NCC) - NC fed to Eimeria spp. challenged birds; BD with 0.2% A. subrufescens inclusion for challenged birds (As), BD with 0.2% P. ostreatus inclusion for challenged birds (Po); and a positive control - BD with anticoccidial and antibiotic inclusion for challenged birds (ATB).3. At 11 d.o., the birds were each inoculated orally with 1 ml solution containing 2 × 105 sporulated oocysts/ml Eimeria acervulina and 2 × 104 sporulated oocysts/ml E. maxima and E. tenella.4. Birds subjected to Eimeria spp. challenge up to 21 d of age had greater crypt depth, indicating that the presence of undesirable microorganisms had an effect on cell proliferation.5. At 21 d old, the birds receiving ATB had higher average weight gain (AWG), feed intake (AFI), and feed conversion ratio (FCR) compared to those fed diets supplemented with mushrooms (As or Po). For the total rearing period (42 days), the birds that received ATB had higher AWG and AFI (P < 0.001) compared to those that received As or Po diets. Feeding avilamycin did not affect (P = 0.0676) FCR compared to the As or Po diet groups.6. From the morphometric and blood analyses there were no differences between broilers fed ATB, Po or As diets in either rearing periods. However, Po and As supplementation lowered blood triglyceride levels. At 21d there was a difference (P < 0.05) for MCV and haemoglobin, in which the mushrooms were similar to the antibiotic. At 42 d, there was a difference (P < 0.05) in haematocrit, erythrocyte, MCV, H: L, protein and albumin variables, in which the use of mushrooms was similar to the positive control, demonstrating that both (mushrooms and antibiotics) promoted a certain improvement in the health of the chickens.7. A. subrufescens and P. ostreatus can be used in broiler diets without compromising intestinal or haematological status, however, these ingredients did not result in improvements in performance.

A multiplex PCR assay for the simultaneous detection and discrimination of the seven Eimeria species that infect domestic fowl.

This study reports the development of a novel multiplex PCR assay based on SCAR (Sequence-Characterised Amplified Region) markers for the simultaneous diagnosis of the 7 Eimeria species that infect domestic fowl. Primer pairs specific for each species were designed in order to generate a ladder of amplification products ranging from 200 to 811 bp. Sensitivity tests for each species were carried out, showing a detection threshold of 1-5 pg, which corresponds approximately to 2-8 sporulated oocysts. Distinct isolates of the 7 Eimeria species from different geographical sources were tested and successfully detected by the assay. All the species were amplified homogeneously, whether or not one of them was present in a high quantity, indicating that there was no cross-interference. The assay was also tested with different sources of Taq DNA polymerase and thermocycler models, confirming the high reproducibility of the reaction. The economy of consumables and labour represented by a single-tube reaction greatly facilitates the molecular diagnosis of a large number of samples, making it appropriate for field epizootiological surveys. We propose the use of this multiplex PCR assay as a rapid and cost-effective diagnostic method for the detection and discrimination of the 7 Eimeria species that infect domestic fowl.

A survey of the inter- and intraspecific RAPD markers of Eimeria spp. of the domestic fowl and the development of reliable diagnostic tools.

Coccidiosis of domestic fowl is a protozoan disease, caused by seven distinct species of the genus Eimeria, which is responsible for important economic losses in poultry production. In order to select RAPD primers for the discrimination of these seven Eimeria species, we carried out an initial screening using samples of E. acervulina, E. tenella and E. maxima. Out of 150 primers tested, 110 generated band profiles specific for each one of these species. A subset of 14 oligonucleotides were also tested for the simultaneous differentiation of the seven species, resulting in 11 discriminative primers. The intraspecific discrimination was assessed for five different species, using samples from different geographic regions including three continents. Numerous primers exhibited highly discriminative band profiles containing strain-specific markers, with a higher variability being observed among strains of E. acervulina than among E. tenella and E. maximastrains. However, no major differences were observed in the band patterns from strains collected in locations near to one another compared to strains originating from distantly located regions. Because RAPD is a technique performed under low stringency conditions, it suffers from poor reproducibility. Aiming at obtaining more reliable markers that might be universally used, we started an effort to convert species-specific RAPD fragments into SCAR markers. An initial conversion of 25 RAPD markers into SCARs, followed by validation of their specificity, resulted in 14 totally new Eimeria species-specific markers that can be used for the molecular diagnosis of the seven species that infect domestic fowl. This work represents a first step in the development of a set of species-specific SCARs that will be useful as tools for molecular diagnosis, genome mapping, and genetic diversity studies.

Genealogical analyses of rabies virus strains from Brazil based on N gene alleles.

Thirty rabies virus isolates from cows and vampire bats from different regions of São Paulo State, Southeastern Brazil and three rabies vaccines were studied genetically. The analysis was based on direct sequencing of PCR-amplified products of 600 nucleotides coding for the amino terminus of nucleoprotein gene. The sequences were checked to verify their genealogical and evolutionary relationships and possible implication for health programmes. Statistical data indicated that there were no significant genetic differences between samples isolated from distinct hosts, from different geographical regions and between samples collected in the last two decades. According to the HKA test, the variability observed in the sequences is probably due to genetic drift. Since changes in genetic material may produce modifications in the protein responsible for immunogenicity of virus, which may eventually cause vaccine failure in herds, we suggest that continuous efforts in monitoring genetic diversity in rabies virus field strains, in relation to vaccine strains, must be conducted.

Comparison of the genomes of two Xanthomonas pathogens with differing host specificities.

The genus Xanthomonas is a diverse and economically important group of bacterial phytopathogens, belonging to the gamma-subdivision of the Proteobacteria. Xanthomonas axonopodis pv. citri (Xac) causes citrus canker, which affects most commercial citrus cultivars, resulting in significant losses worldwide. Symptoms include canker lesions, leading to abscission of fruit and leaves and general tree decline. Xanthomonas campestris pv. campestris (Xcc) causes black rot, which affects crucifers such as Brassica and Arabidopsis. Symptoms include marginal leaf chlorosis and darkening of vascular tissue, accompanied by extensive wilting and necrosis. Xanthomonas campestris pv. campestris is grown commercially to produce the exopolysaccharide xanthan gum, which is used as a viscosifying and stabilizing agent in many industries. Here we report and compare the complete genome sequences of Xac and Xcc. Their distinct disease phenotypes and host ranges belie a high degree of similarity at the genomic level. More than 80% of genes are shared, and gene order is conserved along most of their respective chromosomes. We identified several groups of strain-specific genes, and on the basis of these groups we propose mechanisms that may explain the differing host specificities and pathogenic processes.

Exsudaçäo celular induzida pela carragenina na cavidade peritoneal de pintos / Cellular exudation induced by carrageenan in the peritoneal cavity of chicks

A resposta celular, induzida pela injeçäo de carragenina (4,5 ml de soluçäo de Ringer-Locke a 0,2 por cento w/v) na cavidade peritoneal de pintos (machos Isa Brown de três a seis semanas de idade), foi avaliada pela contagem total e diferencial das células inflamatórias de exsudatos colhidos de 0,5 a 4 horas pós-estímulo. Foi observado ao longo de 1 a 4 horas após a injúria um aumento da exsudaçäo leucocitária, com predominância de heterófilos. Macrófagos foram o segundo tipo celular predominante, seguidos por linfócitos, eosinófilos e basófilos, em ordem decrescente. Trombócitos näo foram observados em número significante no exsudato inflamatório. Os resultados apresentados neste trabalho indicam que a cavidade peritoneal pode ser usada como um bom modelo para o estudo da resposta inflamatória aguda em galinhas

Effect of different vehicles in carrageenan suspension on the rate of the inflammatory response of chicks

Este trabalho descreve o padräo de resposta de edema, aumento de permeabilidade vascular e exsudaçäo celular induzidos pela injeçäo de diferentes suspensöes de carragenina nos coxins plantares de 80 pintos machos, de três a quatro semanas de idade. As suspensöes de carragenina 0,5 por cento foram preparadas em: soluçäo de Ringer-Locke (RL), soluçäo aquosa de Glicose (G), água desmineralizada (W) ou tampäo fosfato salino (PBS). Antes, e às 0:15, 0:30, 1:00, 1:30, 2:00, 2:30, 3:00, 3:30 e 4:00 horas após a injúria, o volume da pata e a permeabilidade vascular foram avaliados através de pletismografia e extravasamento de Azul de Evans respectivamente. A exsudaçäo celular foi observada em cortes finos de tecido corado, 0:30, 1:30, 2:30 e 4:00 horas após a injeçäo de carragenina ou somente do veículo. A resposta inflamatória variou de acordo com a suspensäo de carragenina utilizada. A suspensäo C/PBS induziu uma resposta inflamatória menos intensa nos coxins plantares do que as suspensöes de C/W, C/G e C/RL