Evaluation of the contribution of the placentomal fusion during gestation in cattle

The placenta of mammals is a structure formed by the juxtaposition of the fetal membranes and the maternal tissues. The main function of the placenta is to regulate the physiological interchange between the fetus and the mother as well as to operate as an important endocrine organ during the gestation. The placentomal fusions were characterized throughout gestation of cattle using macroscopic, histological and flow cytometry analyses. Analyzing the cell cycle phases with a flow cytometry, a balance between the G2M phase and apoptosis was observed, suggesting that the placentomal fusions do not interfere in the placentary maturation process, which is a pre-requirement for the fetal-maternal disconnection and the release of fetal membrane. (C) 2011 Elsevier B.V. All rights reserved.

Loss of Methylation at H19 DMD Is Associated with Biallelic Expression and Reduced Development in Cattle Derived by Somatic Cell Nuclear Transfer

Although cloning of mammals has been achieved successfully, the percentage of live offspring is very low because of reduced fetal size and fewer implantation sites. Recent studies have attributed such pathological conditions to abnormal reprogramming of the donor cell used for cloning. The inability of the oocyte to fully restore the differentiated status of a somatic cell to its pluripotent and undifferentiated state is normally evidenced by aberrant DNA methylation patterns established throughout the genome during development to blastocyst. These aberrant methylation patterns are associated with abnormal expression of imprinted genes, which among other genes are essential for normal embryo development and gestation. We hypothesized that embryo loss and low implantation rates in cattle derived by somatic cell nuclear transfer (SCNT) are caused by abnormal epigenetic reprogramming of imprinted genes. To verify our hypothesis, we analyzed the parental expression and the differentially methylated domain (DMD) methylation status of the H19 gene. Using a parental-specific analysis, we confirmed for the first time that H19 biallelic expression is tightly associated with a severe demethylation of the paternal H19 DMD in SCNT embryos, suggesting that these epigenetic anomalies to the H19 locus could be directly responsible for the reduced size and low implantation rates of cloned embryos in cattle.

Delivery of cloned offspring: experience in Zebu cattle (Bos indicus)

The production of a healthy cloned calf is dependent on a multitude of successful steps, including reprogramming mediated by the oocyte, the development of a functional placenta, adequate maternal-fetal interaction, the establishment of a physiological metabolic setting and the formation of a complete set of well-differentiated cells that will eventually result in well-characterised and fully competent tissues and organs. Although the efficiency of nuclear transfer has improved significantly since the first report of a somatic cell nuclear transfer-derived animal, there are many descriptions of anomalies concerning cloned calves leading to high perinatal morbidity and mortality. The present article discusses some our experience regarding perinatal and neonatal procedures for cloned Zebu cattle (B. indicus) that has led to improved survival rates in Nellore cloned calves following the application of such `labour-intensive technology`.

Embryo Mitochondrial DNA Depletion Is Reversed During Early Embryogenesis in Cattle

The extensive replication of mitochondria during oogenesis and the wide variability in mitochondrial DNA ( mtDNA) copy numbers present in fully grown oocytes indicate that mtDNA amount may play an important role during early embryogenesis. Using bovine oocytes derived from follicles of different sizes to study the influence of mtDNA content on development, we showed that oocytes obtained from small follicles, known to be less competent in developing into blastocysts, contain less mtDNA than those originating from larger follicles. However, because of the high variability in copy number, a more accurate approach was examined in which parthenogenetic one-cell embryos were biopsied to measure their mtDNA content and then cultured to assess development capacity. Contrasting with previous findings, mtDNA copy number in biopsies was not different between competent and incompetent embryos, indicating that mtDNA content is not related to early developmental competence. To further examine the importance of mtDNA on development, one-cell embryos were partially depleted of their mtDNA (64% +/- 4.1% less) by centrifugation followed by the removal of the mitochondrial-enriched cytoplasmic fraction. Surprisingly, depleted embryos developed normally into blastocysts, which contained mtDNA copy numbers similar to nonmanipulated controls. Development in depleted embryos was accompanied by an increase in the expression of genes (TFAM and NRF1) controlling mtDNA replication and transcription, indicating an intrinsic ability to restore the content of mtDNA at the blastocyst stage. Therefore, we concluded that competent bovine embryos are able to regulate their mtDNA content at the blastocyst stage regardless of the copy numbers accumulated during oogenesis.

High genetic diversity in field isolates of Trypanosoma theileri assessed by analysis of cathepsin L-like sequences disclosed multiple and new genotypes infecting cattle in Thailand

In this study, we describe the first survey in Thailand of Trypanosoma theileri, a widespread and prevalent parasite of cattle that is transmitted by tabanid flies. Investigation of 210 bovine blood samples of Thai cattle from six farms by hematocrit centrifuge technique (HCT) revealed 14 samples with trypanosomes morphologically compatible to T. theileri. Additional animals were positive for T. theileri by PCR based on the Cathepsin L-like sequence (TthCATL-PCR) despite negative by HCT, indicating cryptic infections. Results revealed a prevalence of 26 +/- 15% (95% CI) of T. theileri infection. Additionally, 12 samples positive for T. theileri were detected in cattle from other 11 farms. From a total of 30 blood samples positive by HCT and/or PCR from 17 farms, seven were characterized to evaluate the genetic polymorphism of T. theileri through sequence analysis of PCR-amplified CATL DNA sequences. All CATL sequences of T. theileri from Thai cattle clustered with sequences of the previously described phylogenetic lineages TthI and TthII, supporting only two major lineages of T. theileri in cattle around the world. However, 11 of the 29 CATL sequences analyzed showed to be different, disclosing an unexpectedly large polymorphic genetic repertoire, with multiple genotypes of T. theileri not previously described in other countries circulating in Thai cattle. (C) 2011 Elsevier B.V. All rights reserved.

Characterization of spliced leader genes of Trypanosoma (Megatrypanum) theileri: phylogeographical analysis of Brazilian isolates from cattle supports spatial clustering of genotypes and parity with ribosomal markers

Trypanosoma (Megatrypanum) theileri from cattle and trypanosomes of other artiodactyls form a clade of closely related species in analyses using ribosomal sequences. Analysis of polymorphic sequences of a larger number of trypanosomes from broader geographical origins is required to evaluate the Clustering of isolates as suggested by previous studies. Here, we determined the sequences of the spliced leader (SL) genes of 21 isolates from cattle and 2 from water buffalo from distant regions of Brazil. Analysis of SL gene repeats revealed that the 5S rRNA gene is inserted within the intergenic region. Phylogeographical patterns inferred using SL sequences showed at least 5 major genotypes of T. theileri distributed in 2 strongly divergent lineages. Lineage TthI comprises genotypes IA and IB from buffalo and cattle, respectively, from the Southeast and Central regions, whereas genotype IC is restricted to cattle from the Southern region. Lineage Tth II includes cattle genotypes IIA, which is restricted to the North and Northeast, and IIB, found in the Centre, West, North and Northeast. PCR-RFLP of SL genes revealed valuable markers for genotyping T. theileri. The results of this study emphasize the genetic complexity and corroborate the geographical structuring of T. theileri genotypes found in cattle.

Structure and Mode of Action of Microplusin, a Copper II-chelating Antimicrobial Peptide from the Cattle Tick Rhipicephalus (Boophilus) microplus

Microplusin, a Rhipicephalus (Boophilus) microplus antimicrobial peptide (AMP) is the first fully characterized member of a new family of cysteine-rich AMPs with histidine-rich regions at the N and C termini. In the tick, microplusin belongs to the arsenal of innate defense molecules active against bacteria and fungi. Here we describe the NMR solution structure of microplusin and demonstrate that the protein binds copper II and iron II. Structured as a single alpha-helical globular domain, microplusin consists of five alpha-helices: alpha 1 (residues Gly-9 to Arg-21), alpha 2 (residues Glu-27 to Asn-40), alpha 3 (residues Arg-44 to Thr-54), alpha 4 (residues Leu-57 to Tyr-64), and alpha 5 (residues Asn-67 to Cys-80). The N and C termini are disordered. This structure is unlike any other AMP structures described to date. We also used NMR spectroscopy to map the copper binding region on microplusin. Finally, using the Gram-positive bacteria Micrococcus luteus as a model, we studied of mode of action of microplusin. Microplusin has a bacteriostatic effect and does not permeabilize the bacterial membrane. Because microplusin binds metals, we tested whether this was related to its antimicrobial activity. We found that the bacteriostatic effect of microplusin was fully reversed by supplementation of culture media with copper II but not iron II. We also demonstrated that microplusin affects M. luteus respiration, a copper-dependent process. Thus, we conclude that the antibacterial effect of microplusin is due to its ability to bind and sequester copper II.

Diagnoses of fipronil resistance in Brazilian cattle ticks (Rhipicephalus (Boophilus) microplus) using in vitro larval bioassays

Fipronil is a phenylpyrazolic insecticide that is widely used in agriculture and has been recently used to control the cattle tick, Rhipicephalus (Boophilus) microplus. Because of the serious problems associated with resistance to the available acaricides, this product has been used as an important alternative to control acaricide-resistant ticks. The objective of this work was to analyse the fipronil sensitivity of ticks that were collected from farms with a history of fipronil use by larval bioassays. A total of 11 Brazilian tick populations were studied: one population from Rio Grande do Sul, one population from Mato Grosso do Sul and nine populations from Sao Paulo. To validate the assays, susceptible reference strains, POA (Porto Alegre, Brazil) and Mozo (Dilave, Uruguay), and ticks from six different farms that never used fipronil were tested. The resistance of various tick populations to technical grade fipronil (95.3%) was primarily evaluated using the larvae immersion test (LIT) and the larval packet test (LPT), when a sufficient number of larvae was collected. Using the LIT, the resistance ratios (RR(50)) of the tick populations from Rio Grande do Sul and Mato Grosso do Sul were 14.9 and 2.6, respectively, and the populations derived from Sao Paulo had RR(50)s ranging from 2.5 to 6.9. Four populations were evaluated with the LPT, and two populations displayed lower RR(50), while other populations displayed higher RR(50) than those determined by the LIT. This article reports the first cases of fipronil resistance in Brazil and highlights the LIT as a more sensitive technique for the evaluation of fipronil resistance in R. (B) microplus ticks. We suggest the use of the LIT as an evaluation tool for monitoring fipronil resistance in the control programmes of R. (B) microplus. (C) 2010 Elsevier B.V. All rights reserved.

Estimates of genetic parameters for visual scores and their correlation with production and reproductive traits in Brahman cattle

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Anthelmintic efficacy of oral trichlorfon solution against ivermectin resistant nematode strains in cattle

Infected calves from two different rural estates in Brazil were studied to assess the anthelmintic efficacy of oral trichlorfon against naturally occurring ivermectin resistant parasitic nematode strains. In experiment 1, infected animals were from a region where ivermectin resistant populations of Haemoncus placei, Cooperia punctata,Cooperia spatulata and Trichuris discolor have recently been identified. Six calves with natural gastrointestinal nematode infections were treated with 48.5 mg/kg aqueous trichlorfon administered orally and six calves acted as a non-treated control group. In experiment 11 24 naturally infected calves were selected to enter one of four treatment groups, six animals each received: 48.5 mg/kg oral trichlorfon; 200 mu g/kg subcutaneous 1% ivermectin; 630 mu g/kg subcutaneous 3.15% ivermectin; or no treatment (control group). Gastrointestinal helminths were counted and identified post-mortem at 7 days (trichlorfon and 1% ivermectin treated and untreated animals) or 14 days (3.15% ivermectin treated and untreated animals) after administration of the test agents. Experiment I identified a high level efficacy for oral trichlorfon against four helminth species that have previously been shown to be ivermectin resistant in this geographical region: percentage efficacy was 99.82% against adult H. placei, 99.18% against C. punctata, 99.33% against C. spatulata, 81.06% against T. axei, 98.46% against Oesophagostomum radiatum and 100% against T. discolor. Trichlorfon also showed activity against the ivermectin (1% and 3.15%) resistant helminth species identified in experiment 11, attaining efficacy levels of 99.17% against H. placei, 98.46% against C punctata and 100.00% against T. discolor. These findings indicate that oral trichlorfon is an effective treatment option in the management of cattle infected with ivermectin resistant helminths. (C) 2009 Elsevier B.V. All rights reserved.

Preferential infection sites of Cysticercus bovis in cattle experimentally infected with Taenia saginata eggs

The preferential sites of infection of Cysticercus bovis were evaluated in the skeletal muscle and entrails of 25 cattle that were experimentally infected with Taenia saginata (2 x 10(4) eggs). Two other animals were not inoculated (control). Ninety days after inoculation, all the cattle were euthanized. The carcasses were deboned and dissected into 26 anatomical sections (masseter muscles, brain, tongue, esophagus, heart, diaphragm, lungs, liver, kidneys, spleen, top sirloin butt, bottom sirloin butt, outside round, top (inside) round, transversus abdominus, top sirloin cap, strip loin, full tenderloin, eye of round, knuckle, shoulder clod, foreshank, shank, chuck, back ribs, and tail muscles). The dissected tissues were sliced into 5 mm sections. From the 25 cattle, 9258 C. bovis (cysticerci) were recovered; 75.02% (6946) of these were recovered from skeletal muscles and 24.98% (2312) from the entrails. A high parasitism level was found in the shoulder clod (12.55%), heart (11.02%), liver (9.48%), masseter muscles (8.51%), chuck (8.25%), strip loin and full tenderloin (7.26%), knuckle (6.63%), and back ribs (5.53%), totaling 69.23% (5738) of all of the detected cysticerci. on the other hand, there was a low C. bovis parasitism level in the brain, spleen, tail muscles, kidneys, esophagus, and diaphragm, representing just 3.9% of the total number of cysticerci. Given these results, we conclude that specific skeletal musculature regions, such as the shoulder blade, chuck, strip loin and full tenderloin, knuckle, back ribs and top round, which are not officially examined in many countries, are effective sites to efficiently screen C. bovis infection. To date, these regions have not been considered as preferential sites of C. bovis infection. Based on our work, however, these regions deserve greater attention from health inspectors because they contained a greater number of Cysticercus than the other regions of carcasses that are parasitized by T. saginata larvae. (c) 2010 Elsevier Ltd. All rights reserved.

Toxoplasma gondii: Infection natural congenital in cattle and an experimental inoculation of gestating cows with oocysts

Two studies, of a natural infection and an experimental infection, were performed in order to study congenital transmission of Toxoplasma gondii in cattle. In the first study, 50 fetuses were harvested from gestating cows that were eutanasied at a municipal slaughterhouse in Jaboticabal, São Paulo state, Brazil. In the second study, 11 gestating cows were divided into four groups for inoculation with T. gondii: GI consisted of three cows inoculated with 1.0 x 10(5) oocysts during their first trimester of gestation; GII consisted of three cows inoculated with 1.0 x 10(5) oocysts during their second trimester of gestation; GIII consisted of three cows inoculated with 1.0 x 10(5) oocysts during their last trimester of gestation; and GIV consisted of two control cows, one during its first and the other during its second trimester of gestation. In both studies, the presence of T. gondii was confirmed both indirectly by immunofluorescence assay (IFAT). In the natural infection experiment, 18% (9/50) of the gestating cows were confirmed to have specific antibodies (IFAT - 1:64) against T. gondii. The bioassay was able to diagnose the presence of T. gondii in the tissue samples from three calves. In the second experiment, the nine cows from groups I, II and III presented with specific antibodies (IFAT) against T. gondii. In contrast, T. gondii could not be detected by IFAT, histopathological examination or the bioassay in any of the nine calves born to cows experimentally infected with T. gondii oocysts. Based on the results from both studies, we conclude that congenital infection of T. gondii in cattle, while infrequent, does occur naturally. The pathogenicity of the strain of T. gondii may influence the likelihood of this route of transmission. (C) 2010 Elsevier B.V. All rights reserved.

Organic matter fractions and soil fertility under the influence of liming, vermicompost and cattle manure

Os efeitos do vermicomposto de esterco de curral associado à calagem em atributos da fertilidade do solo foram avaliados através de experimento em vasos empregando um Latossolo Vermelho, distrófico, textura média. Cinco doses do vermicomposto (equivalentes a 0; 28; 42; 56 e 70 t ha-1, peso seco) e cinco doses de calcário (visando elevar a saturação por bases a: 20; 30; 40; 50 e 60%) foram combinadas em esquema fatorial, sendo as amostras de solo incubadas por 180 dias. Para comparação entre o vermicomposto e o esterco de curral, amostras do mesmo solo receberam o equivalente a 70 t ha-1 do esterco de curral que originou o vermicomposto e as cinco doses de calcário listadas anteriormente. Através do cálculo do Índice de Eficiência Agronômica, foi verificado que o potencial de fornecimento de K e de Mg pelo esterco é maior do que o do vermicomposto, e que o de P, é semelhante. O vermicomposto aumentou os teores de Ca2+ e de matéria orgânica (MO), os valores de pH em CaCl2 e a CTC a pH 7. Com o aumento das doses de vermicomposto houve diminuição do C-ácidos húmicos e aumento do C-humina e com a calagem o C-total não aumentou mas houve diminuição do C-ácidos húmicos.

Genetic associations between weight at maturity and maturation rate with ages and weights at first and second calving in Canchim beef cattle

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Association of an insulin-like growth factor 1 gene microsatellite with phenotypic variation and estimated breeding values of growth traits in Canchim cattle

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)