RESUMO
The objective of the present study was to evaluate the effect of estradiol valerate administered at the beginning of the ovulation synchronization protocol on the pregnancy rate of Bos indicus cows. In the experiments, the following products from MSD, Sao Paulo, Brazil were used: estradiol valerate (EV), estradiol benzoate (EB), intravaginal progesterone device (P4), estradiol cypionate (EC), equine chorionic gonadotropin (eCG) and cloprostenol (PGF). In Experiment 1, Bos indicus cows (n=899) with a body condition score (BCS) of 2.76 ± 0.01 were included in a 3 (device) × 2 (protocol: 5 mg of EV or 2 mg of EB) factorial design. There were three types of P4 devices: a new device (New), a device previously used for 9 days (1×), and a device previously used for 18 days (2×). Nine days later (D9), the P4 device was removed, and cows received 300 IU of eCG. In addition, cows in the EB group received 1 mg of EC and 265 µg of PGF. Timed artificial insemination (TAI) was performed 48 h after P4 device removal in the EB group (TAI48) and 54 h after P4 device removal in the EV group (TAI54). In Experiment 2, Bos indicus cows (n=434) with a BCS of 2.62 ± 0.01 received a new P4 device or one previously used for 9 days and 5 mg of EV. On D9, all cows received 300 IU of eCG, and the P4 devices were removed and distributed in TAI48 and TAI54 cows. In Experiment 3, Bos indicus cows (n=429) with a BCS of 2.80 ± 0.01 were divided into the control and EV/EC groups. All cows received a P4 device. In addition, cows in the control group received 2 mg of EB, and cows in the EV/EC group received 5 mg of EV on D0. On D9, all cows received 1 mg of EC and 300 IU of eCG, and the P4 devices were removed. Cows in the control group also received 265 µg of PGF. All cows were inseminated 48 h after the removal of the P4 device. In Experiment 1, there was no effect of the interaction between protocol and P4 device on the occurrence of estrus (P=0.45) or on the pregnancy per artificial insemination ratio (P/AI; P=0.30). In addition, the occurrence of estrus and P/AI were not different between in the two estradiol groups (P=0.12 and P=0.82) and across the types of intravaginal P4 device (P=0.91 and P=0.47). In Experiment 2, the pregnancy rate was lower (tendency) in TAI48 cows (P=0.07). In Experiment 3, the estrus rate (P=0.12) and P/AI (P=0.56) were similar between the experimental groups. In summary, protocols using estradiol valerate without exogenous ovulation induction require adjustments in the timing of AI from 48 to 54 h after P4 device removal. However, a combination of estradiol valerate at the beginning of the protocol and estradiol cypionate nine days later successfully induced ovulation in Bos indicus cows inseminated 48 h after P4 device removal.
Assuntos
Estradiol , Estradiol/análogos & derivados , Progesterona , Gravidez , Feminino , Bovinos , Animais , Cavalos , Brasil , Estradiol/farmacologia , Progesterona/farmacologia , Ovulação , Inseminação Artificial/veterinária , Inseminação Artificial/métodos , Sincronização do Estro/métodosRESUMO
BACKGROUND: Neonatal calf diarrhea (NCD) is the leading cause of calf morbidity and mortality in beef cattle. Cow's vaccination in last stage of pregnancy is one of the most important measures to mitigate the risk of NCD outbreaks. The aim of this study was to evaluate the efficacy of prepartum single dose vaccination against NCD, especially Bovine Rotavirus type A (BoRVA) and Bovine Coronavirus (BCoV), in Nelore dams and offspring. A total of 117 pregnant cows (n = 81) and heifers (n = 36) were distributed in two groups, vaccinated (VAC: cows = 40; heifers = 19) and non-vaccinated (NVAC: cows = 41; heifers = 17). Vaccination occurred between 60 to 50 days before the expected calving date with a single dose of a water-in-oil (W/O) vaccine, and NVAC group received a dose of saline solution 0.9%. Blood samples were collected before vaccination and 30 days after to evaluate the antibody (Ab) response. Specific IgG1 Abs against BoRVA and BCoV were measured by using an Enzyme Linked Immuno Sorbent Assay (ELISA). Calves' births were monitored, and the transference of passive immunity was evaluated. Diarrhea was monitored in the first 30 days of age, and fecal samples were collected for identification of the etiological agent. RESULTS: Higher titers of IgG1 Ab against BoRVA and BCoV was observed in the VAC group than NVAC group in the cow (P < 0.0001) and total dams categories (P < 0.0001). The titer of specific IgG1 Abs in the calves' serum reflected the dams response, observing higher IgG1 Ab titers for BoRVA (P < 0.0016) and BCoV (P < 0.0095) in the offspring born to VAC cows and higher IgG1 Ab titers for BoRVA(P < 0.0171) and BCoV (P < 0.0200) in the offspring born to VAC total dams. The general incidence of diarrhea observed was 18.6% (11/59) and 29.3% (17/58) in the calves born to the VAC and NVAC group, respectively. CONCLUSIONS: Prepartum vaccination with a single dose of the vaccine tested increased the titers of IgG1 Ab against BCoV and BoRVA, and it could be used as a preventive strategy to decrease the NCD occurrence in Nelore calves.
Assuntos
Doenças dos Bovinos , Doenças não Transmissíveis , Animais , Bovinos , Diarreia/prevenção & controle , Diarreia/veterinária , Feminino , Imunoglobulina G , Gravidez , Vacinação/veterináriaRESUMO
Cattle are an important source of zoonotic species of Cryptosporidium for humans. The aim of this study was to investigate the presence of Cryptosporidium, identify the species and determine the risk factors relating to environment, animals and management among dairy calves in eight Brazilian states. A total of 408 fecal samples from calves aged 1-60 days were analyzed. An epidemiological questionnaire was completed. Sample screening was performed using Ziehl-Neelsen technique and the positive samples were subjected to nested PCR. Cryptosporidium species were identified by means of the PCR-RFLP technique, using SSPI, ASEI and MBOII enzymes. The Ziehl-Neelsen technique showed that 89.7% (35/39) of the farms and 52.9% (216/408) of the samples were positive. Through nested PCR, these protozoa were detected in 54.6% of the samples. The 56 samples subjected to PCR-RFLP presented Cryptosporidium parvum. There was higher prevalence of the parasite in animals aged 7 to 28 days (62.6%). Diarrhea, ages between seven and 28 days and a spring water source were factors associated with the risk of infection. The calf hutch-type management system was associated with reduced infection. These findings demonstrate the high level of Cryptosporidium spp. circulation in cattle herds and the predominance of the species C. parvum.
Assuntos
Doenças dos Bovinos , Criptosporidiose , Cryptosporidium , Animais , Brasil/epidemiologia , Bovinos , Doenças dos Bovinos/diagnóstico , Doenças dos Bovinos/epidemiologia , Criptosporidiose/epidemiologia , Cryptosporidium/genética , Fazendas , Fezes , PrevalênciaRESUMO
Abstract Cattle are an important source of zoonotic species of Cryptosporidium for humans. The aim of this study was to investigate the presence of Cryptosporidium, identify the species and determine the risk factors relating to environment, animals and management among dairy calves in eight Brazilian states. A total of 408 fecal samples from calves aged 1-60 days were analyzed. An epidemiological questionnaire was completed. Sample screening was performed using Ziehl-Neelsen technique and the positive samples were subjected to nested PCR. Cryptosporidium species were identified by means of the PCR-RFLP technique, using SSPI, ASEI and MBOII enzymes. The Ziehl-Neelsen technique showed that 89.7% (35/39) of the farms and 52.9% (216/408) of the samples were positive. Through nested PCR, these protozoa were detected in 54.6% of the samples. The 56 samples subjected to PCR-RFLP presented Cryptosporidium parvum. There was higher prevalence of the parasite in animals aged 7 to 28 days (62.6%). Diarrhea, ages between seven and 28 days and a spring water source were factors associated with the risk of infection. The calf hutch-type management system was associated with reduced infection. These findings demonstrate the high level of Cryptosporidium spp. circulation in cattle herds and the predominance of the species C. parvum.
Resumo O gado é uma fonte importante de espécies zoonóticas de Cryptosporidium para o homem. O objetivo deste estudo foi investigar a presença de Cryptosporidium, identificar a espécie e determinar os fatores de risco relacionados ao meio ambiente, aos animais e ao manejo em bezerros leiteiros em oito estados brasileiros. Um total de 408 amostras fecais de bezerros, com idade entre 1 e 60 dias, foram analisadas. Um questionário epidemiológico foi preenchido. A triagem das amostras foi realizada pela técnica de Ziehl-Neelsen, e as amostras positivas foram submetidas à "nested" PCR. As espécies de Cryptosporidium foram identificadas pela técnica de PCR-RFLP, utilizando-se as enzimas SSPI, ASEI e MBOII. A técnica de Ziehl-Neelsen mostrou que 89,7% (35/39) das fazendas e 52,9% (216/408) das amostras foram positivas. Por meio de nested PCR, esses protozoários foram detectados em 54,6% das amostras. As 56 amostras submetidas à PCR-RFLP apresentaram Cryptosporidium parvum. Houve maior prevalência do parasita em animais de 7 a 28 dias (62,6%). Diarreia, idade entre sete e 28 dias, e fonte de água mineral foram fatores associados ao risco de infecção. O sistema de manejo do tipo "casinha" para bezerros foi associado à redução da infecção. Esses achados demonstram o alto nível de Cryptosporidium spp. em circulação nos rebanhos bovinos e o predomínio da espécie C. parvum.
Assuntos
Animais , Doenças dos Bovinos/diagnóstico , Doenças dos Bovinos/epidemiologia , Criptosporidiose/genética , Criptosporidiose/epidemiologia , Brasil/epidemiologia , Bovinos , Prevalência , Fezes , FazendasRESUMO
The present study attempted to verify the prevalence of and risk factors for diarrhea-causing agents in dairy calves from Brazil. Additionally, ages with a higher risk of occurrence for each agent were verified by means of the receiver operating characteristic (ROC) curve. The collections were performed on 39 farms, belonging to 29 municipalities located in eight states of Brazil. It was possible to conclude that the prevalence of Coronavirus, Rotavirus, Cryptosporidium spp., Eimeria spp., and nematodes was 7.20% (95% CI 4.54-9.78), 6.37% (95% CI 3.85-8.89), 51.52% (95% CI 45.26-55.57), 3.46% (95% CI 2.24-4.67), and 3.46% (95% CI 2.24-4.67), respectively. Ages with higher probabilities of occurrence of these diseases in calves were < 10, > 8, > 6, > 37, and > 36 days, respectively. Diarrhea occurred more significantly (P < 0.0001) in animals less than 21 days old and mainly on those receiving milk through automatic feeders (P < 0.001). Cryptosporidium spp. were a risk factor for the occurrence of Rotavirus, and vice versa (P = 0.0039) and presented a positive correlation with Coronavirus (P = 0.0089). Calves that drink water from rivers, streams, and ponds had a higher chance of being infected by Eimeria spp. (P < 0.0001), as well as developing infection by nematodes (P < 0.0001). The results found in this study highlight the importance of studying the agents of diarrhea together, once they act as coinfection where the losses triggered for the owners will involve some of these agents simultaneously.
Assuntos
Doenças dos Bovinos/epidemiologia , Infecções por Coronavirus/veterinária , Criptosporidiose/complicações , Diarreia/veterinária , Infecções por Nematoides/veterinária , Infecções por Rotavirus/veterinária , Animais , Brasil/epidemiologia , Bovinos , Doenças dos Bovinos/parasitologia , Doenças dos Bovinos/virologia , Coronavirus/isolamento & purificação , Criptosporidiose/epidemiologia , Cryptosporidium/isolamento & purificação , Diarreia/parasitologia , Diarreia/virologia , Eimeria/isolamento & purificação , Fazendas , Fezes/parasitologia , Fezes/virologia , Feminino , Infecções por Nematoides/complicações , Prevalência , Curva ROC , Fatores de Risco , Rotavirus/isolamento & purificaçãoRESUMO
Worldwide, neonatal diarrhea is one of the most important health issues affecting dairy calves, and rotavirus A (RVA) is one of its primary causes. Among the measures to mitigate the risk of diarrhea outbreaks, cow vaccination stands out as one of the most important. However, the immune pressure resulting from routine vaccination may be able to select specific G and P genotypes in RVA field strains. This study aimed to determine the frequency and intensity of neonatal diarrhea and the incidence of RVA and attempted to monitor the G and P genotypes present in the RVA strains circulating in a high milk yield cattle herd vaccinated with RVA G6P[5] strain. Fecal samples (n = 1220) from 122 Holstein heifer calves between 0-30 days old that were born from RVA-vaccinated cows were collected at 10 different time points, regardless of the presence or absence of diarrhea. The presence of RVA in fecal samples was determined by the polyacrylamide gel electrophoresis (PAGE) technique and confirmed by reverse transcription polymerase chain reaction (RT-PCR). G and P amplicons from 10 RVA-positive fecal samples from calves of different ages and collections were subjected to nucleotide sequencing. The proportion of the calves and fecal samples that were positive for RVA were 62.3% (76/122) and 8.1% (99/1220), respectively. Using sequence analysis, all 10 RVA field strains presented genotype G10P[11]. The protection of G6P[5] vaccination is clear, as this genotype was not detected in this study, and it is known that vaccination against RVA reduces the incidence of diarrhea independent of genotype involved. This result demonstrates the importance of epidemiological monitoring of RVA genotypes circulating in vaccinated dairy cattle herds to the early detection of new potential pathogenic RVA strains.
Assuntos
Doenças dos Bovinos/virologia , Bovinos/virologia , Infecções por Rotavirus/veterinária , Vacinas contra Rotavirus/administração & dosagem , Rotavirus/genética , Animais , Animais Recém-Nascidos , Doenças dos Bovinos/epidemiologia , Indústria de Laticínios , Diarreia/virologia , Monitoramento Epidemiológico/veterinária , Fezes/virologia , Genótipo , Estudos Longitudinais , Leite , Filogenia , Rotavirus/patogenicidade , Infecções por Rotavirus/epidemiologia , Infecções por Rotavirus/prevenção & controle , Análise de Sequência de DNA , Vacinação/veterináriaRESUMO
Neonatal diarrhea is the main cause of morbidity and mortality in calves up to 30 days old, and rotavirus A (RVA) is the main viral etiology. RVA vaccines are one of the main tools for diarrhea control in neonates. The aim of this cross-sectional study was to monitor by RT-PCR the G and P genotypes of RVA strains identified in dairy cattle herds regularly vaccinated with the RVA UK strain (G6P[5]). Of the 14 randomly selected herds, two were excluded because no calf was diagnosed with diarrhea on the day of fecal collection. Another six herds were also excluded from the study because all 20 diarrheic fecal samples evaluated were RT-PCR-negative. In the remaining six herds, 17 (25.4%) of the 67 diarrheic samples were RVA-positive. One G and P amplicon from each herd were selected for nucleotide sequencing. In the phylogenetic analysis, five RVA strains presented the G6P[11] genotype, and one presented the G10P[11] genotype. The G6 genotype present in all RVA field strains clustered into a distinct phylogenetic arrangement (lineage III) of the UK vaccine strain (lineage IV), characterizing the emergence of a phylogenetically distant G6 strain. In addition, we observed the emergence of strains with G10 and P[11] genotypes characterizing failure in heterologous immune protection. These results show the epidemiological importance of constant monitoring of RVA strains in vaccinated cattle herds and the low frequencies of diarrhea and diagnosis of RVA suggest that a regular vaccination program reduces the frequency and severity of RVA diarrhea in suckling calves.
Assuntos
Doenças dos Bovinos/virologia , Diarreia/veterinária , Infecções por Rotavirus/veterinária , Vacinas contra Rotavirus , Rotavirus/genética , Animais , Animais Recém-Nascidos , Sequência de Bases , Bovinos , Doenças dos Bovinos/prevenção & controle , Estudos Transversais , Diarreia/prevenção & controle , Diarreia/virologia , Fezes/virologia , Genótipo , Filogenia , Infecções por Rotavirus/prevenção & controle , Infecções por Rotavirus/virologiaRESUMO
Oocytes from dairy cattle and buffaloes have severely compromised developmental competence during summer. While analysis of gene expression is a powerful technique for understanding the factors affecting developmental hindrance in oocytes, analysis by real-time reverse transcription PCR (RT-PCR) relies on the correct normalization by reference genes showing stable expression. Furthermore, several studies have found that genes commonly used as reference standards do not behave as expected depending on cell type and experimental design. Hence, it is recommended to evaluate expression stability of candidate reference genes for a specific experimental condition before employing them as internal controls. In acknowledgment of the importance of seasonal effects on oocyte gene expression, the aim of this study was to evaluate the stability of expression levels of ten well-known reference genes (ACTB, GAPDH, GUSB, HIST1H2AG, HPRT1, PPIA, RPL15, SDHA, TBP and YWHAZ) using oocytes collected from different categories of dairy cattle and buffaloes during winter and summer. A normalization factor was provided for cattle (RPL15, PPIA and GUSB) and buffaloes (YWHAZ, GUSB and GAPDH) based on the expression of the three most stable reference genes in each species. Normalization of non-reference target genes by these reference genes was shown to be considerably different from normalization by less stable reference genes, further highlighting the need for careful selection of internal controls. Therefore, due to the high variability of reference genes among experimental groups, we conclude that data normalized by internal controls can be misleading and should be compared to not normalized data or to data normalized by an external control in order to better interpret the biological relevance of gene expression analysis.
Assuntos
Expressão Gênica , Genes Essenciais , Oócitos/metabolismo , Reação em Cadeia da Polimerase em Tempo Real/normas , Reação em Cadeia da Polimerase Via Transcriptase Reversa/normas , Animais , Búfalos , Bovinos , Indústria de Laticínios , Feminino , Perfilação da Expressão Gênica , Oócitos/citologia , Padrões de Referência , Estações do AnoRESUMO
Background: Heat stress (HS) can compromise the female reproductive system, resulting in a massive reduction of reproductive performance. During the period of follicular growth, HS may compromise the oocyte, either because of a direct effect of elevated temperature on the gamete or because of changes in follicular function that would damage oocyte quality. Oocytes harvested from Holstein cows during summer have reduced ability to develop to the blastocyst stage than those harvested during winter. Nevertheless, although there are reports showing a clear effect of HS on oocyte competence to development after IVF, the exact mechanisms by which the oocyte is compromised remain unknown. Review: Resistance to HS is dependent on cow's genotype. Bos indicus breeds (i.e. Gir) are more resistant to elevated temperature and humidity than breeds (i.e. Holstein) that evolved in a temperate climate. This resistant is, among other factors, due to the superior ability of certain breeds to regulate their body temperature. Thus, breed selection is present as a possible tool to minimize the effects of temperature and humidity on milk production and reproduction. On the other hand, Holstein cows are the most common breed used, even in tropical countries (i.e. Brazil), because of their yield in milk production. As a result, there is a considerable effect upon milk production and reproductive performance through the year in tropical and subtropical countries. A possible way to minimize this effect is the employment of reproductive biotechnologies such as fixed-time artificial insemination (FTAI) and embryo transfer (ET). FTAI has been shown to eliminate problems of estrus detection caused by HS. Nevertheless, it is not enough to complete restore herd pregnancy rates because oocytes and early embryos have already been damaged by HS. In contrast, embryos three days after conception orolder are less sensitive to HS. Thus, ET has been used to improve cow's reproductive performance during the hot months of the year (i.e. spring and summer). This is carried out by producing embryos during the fresh months but using them for ET during the HS period. Regarding fertility problems, it is recognized that repeat-breeder (RB) cows cause huge economic problems to the farmer. These cows are characterized by poor fertilization rates and/or early embryonic loss. It is still not certainly known why these cows become RB, when it is the critical period of the year for them, and what should be done to avoid this problem. We have observed that the conception rates in RB Holstein cows were greater after ET (41.7%) than after AI (17.9%). This indicates that ET may be an effective alternative to achieve satisfactory conception rates throughout the year in RB cows, especially during periods of HS. Moreover, when three categories of Holstein cows (heifers, high-producing cows in peak lactation and RB) were subjected to ovum pick up during the summer and the winter, blastocyst rates and quality were significantly affected by both, category and period, being RB cows mostly affected during the summer. Moreover, lower amounts of mitochondrial DNA were found in RB oocytes, suggesting that the low reproductive performance observed in these cows is related to some injure in oocyte quality. Conclusion: HS negatively affects the physiology and fertility of dairy cows and heifers. Concerning about the reproductive field, the effects of HS were detected in vitro and in vivo on oocytes and embryos and also on conception rates. Moreover, RB Holstein cows seem to be even more sensitive to HS and this can probably be related to their oocyte potential into develop in blastocysts. Therefore, efforts should be taken in order to improve the environmental conditions and heat resistance of dairy cows in tropical and subtropical areas. Studies are been conducted to understand the molecular level by which HS disrupt reproduction aiming to develop methods to attenuate or reverse HS consequences upon fertility of dairy cows.