Fabiellen Cristina Pereira
MANEJO DE PASTAGENS E USO DA TÉCNICA PARA DETERMINAR EMISSÕES DE METANO: EFEITO NO COMPORTAMENTO DE PASTOREIO E NA QUALIDADE DA
DIETA
Dissertação submetida como requisito final para a obtenção do grau de mestre em Agroecossistemas pela Universidade Federal de Santa Catarina e mestre em Sistemas de produção animal pela Pontifícia Universidad Católica de Chile em regime de cotutela.
Orientadores: Prof. Dr. Luiz Carlos Pinheiro Machado Filho
Prof. Dr. Daniel Enríquez Hidalgo
Florianópolis/Santiago de Chile 2019
Fabiellen Cristina Pereira
MANEJO DE PRADERAS Y USO DE LA TÉCNICA PARA DETERMINAR EMISIONES DE METANO: EFECTO EN EL COMPORTAMIENTO DE PASTOREO Y LA CALIDAD DE LA
DIETA
Tesis presentada como requisito final para obtener el grado de magíster en Agroecossistemas por la Universidade Federal de Santa Catarina y magíster en Sistemas de producción animal por la Pontifícia Universidad Católica de Chile en regimén de doble grado. Supervisores: Prof. Dr. Luiz Carlos Pinheiro Machado Filho
Prof. Dr. Daniel Enríquez Hidalgo
Florianópolis/Santiago de Chile 2019
Ficha de identificação da obra elaborada pelo autor,
através do Programa de Geração Automática da Biblioteca Universitária da UFSC.
Pereira, Fabiellen Cristina
MANEJO DE PASTAGENS E USO DA TÉCNICA PARA DETERMINAR EMISSÕES DE METANO: EFEITO NO
COMPORTAMENTO DE PASTOREIO E NA QUALIDADE DA DIETA / Fabiellen Cristina Pereira ; orientador, Luiz Carlos Pinheiro Machado Filho, orientador, Daniel Enríquez Hidalgo, 2019.
112 p.
Dissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências Agrárias, Programa de Pós-Graduação em Agroecossistemas, Florianópolis, 2019.
Inclui referências.
Trabalho elaborado em regime de co-tutela.
1. Agroecossistemas. 2. Pastoreio Racional Voisin. 3. Comportamento de bovinos. 4. Produção de gás metano. I. Machado Filho, Luiz Carlos Pinheiro. II. Enríquez Hidalgo, Daniel III. Universidade Federal de Santa Catarina. Programa de Pós-Graduação em Agroecossistemas. IV. Título.
AGRADECIMENTOS
Agradeço à minha família, meus pais e meus irmãos, pelo apoio imensurável durante toda a minha caminhada.
Ao meu orientador Luiz Carlos Pinheiro Machado Filho, pela confiança, pela orientação, por todo o aprendizado e pela amizade construída ao longo de todos esses anos. E ao meu orientador Daniel Enríquez-Hidalgo por ter me oferecido a oportunidade de realizar o mestrado duplo, pelo aprendizado, orientação e amizade.
Aos amigos do Laboratório de Etologia Aplicada por todo o companheirismo, conversas e apoio. E ao laboratório de forragicultura pelo auxílio nas análises.
Aos trabalhadores da Fazenda Experimental da Ressacada e da Fundación AgroUC que me receberam e me auxiliaram, possibilitando a execução das minhas pesquisas.
À Embrapa Cerrados e Embrapa Gado de leite, pesquisadores e funcionários pela oportunidade de realizar as análises fundamentais a minha pesquisa, em especial ao pesquisador Roberto Guimarães Júnior por todo auxílio, ensinamentos e apoio, e ao técnico de laboratório Francisco Bastos por todo o auxílio e amizade.
À Dona Maria e toda sua família por ter me recebido e me acolhido em Planaltina/DF enquanto eu realizei parte da pesquisa na Embrapa. À Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) pela concessão de bolsa de estudo, ao programa de pós-graduação em Agroecossistemas e à Universidade Federal de Santa Catarina e a todos os professores e funcionários que de alguma forma contribuíram para a minha formação pessoal e profissional.
Aos colegas e amigos que se esforçaram para me auxiliar na execução das pesquisas: Amanda Sperandio, Ana Beatriz Almeida Torres, Ana Laura Carrili, Bianca Vandresen, Júlia Vargas, Juliana Butzge, Karolini Tenffen de Sousa, Laura Arias Avilés, Liz Roblez, Luã Veiga, Melissa García Méndez, Macarena Fernandez, Matheus Deniz, Patricia Camila Carrasco, Rodrigo Conceição, Rita Silva e Tasmin Pereda Kihara. E a todos os colegas e amigos que de alguma forma contribuíram com o meu crescimento pessoal e profissional e me apoiaram, dando-me forças para vencer meus obstáculos.
RESUMO
Pastagens melhoradas têm maior valor nutricional, são mais degradáveis no rúmen e proporcionam maior eficiência do comportamento de pastoreio de bovinos. O cultivo de espécies anuais de inverno em consórcio é uma estratégia de melhoramento de pastagens e proporciona benefícios aos animais e ao meio ambiente. O melhoramento deve ser acompanhado por um manejo adequado, respeitando a dinâmica de crescimento da pastagem e permitindo que ela seja pastoreada pelos animais apenas em seu tempo ótimo de repouso. O tempo ótimo de repouso além de indicar a qualidade do pasto, influencia a produção de metano entérico ruminal (CH4). Formas indiretas de mensurar a
produção de CH4 baseiam-se na simulação da fermentação ruminal em
laboratório enquanto técnicas in vivo mensuram a produção emitida diretamente do animal. Entretanto, não se sabe se ao usar técnicas in
vivo os animais alteram seus comportamentos padrões. O primeiro
estudo dessa dissertação objetivou avaliar o efeito da inclusão de ervilhaca (Vicia sativa L.) no cultivo de aveia (Avena strigosa Schreb.), no valor nutricional e na degradabilidade in vitro da pastagem, bem como no comportamento de pastoreio de novilhas. Para tanto realizou-se o cultivo de aveia solteira, aveia em consórcio com ervilhaca e ervilhaca solteira em uma área dividida em 6 blocos, sendo coletadas amostras de pasto para as análises bromatológicas e de degradabilidade in vitro. O comportamento de novilhas foi observado por duas horas e durante esse tempo, foi realizada simulação de pastoreio e a contabilização de taxas de bocado. A inclusão da leguminosa aumentou o teor de proteína da aveia e diminuiu os teores de fibra, promovendo melhor degradabilidade
in vitro e aumento da eficiência de pastoreio das novilhas. O segundo
estudo objetivou avaliar o efeito do tempo de repouso no valor nutricional e degradabilidade da pastagem, e na produção de CH4 in vitro, além da influência no comportamento de pastoreio de novilhas.
Para tanto, três tempos de repouso foram avaliados: 24 dias, 35 dias e 46 dias. Coletas de pasto foram realizadas para análises bromatológicas, de degradabilidade e de produção de CH4 in vitro. Observações do
comportamento de pastoreio ocorreram durante quatro horas, assim como a simulação de pastoreio e a contabilidade de taxas de bocado. A pastagem no tempo de repouso de 24 dias teve melhor valor nutricional e menor produção de CH4. As novilhas selecionaram uma pastagem com
melhor valor nutricional e maior degradabilidade em relação à pastagem ofertada no piquete, mas não houve diferença em relação à produção de
CH4. A taxa de bocados também foi maior no tempo de repouso de 24
dias, mas a frequência de pastoreio foi maior no tempo de repouso de 46 dias. O objetivo do terceiro estudo foi avaliar o efeito da técnica hexafluoreto de enxofre (SF6) para mensurar produção de CH4 in vivo
em bovinos, na produtividade e no comportamento de vacas leiteiras. A produção de leite foi mensurada e amostras de leite foram analisadas para quantificar o rendimento de sólidos. O comportamento das vacas foi avaliado e comparado antes do uso do equipamento, enquanto elas estavam sendo equipadas com o SF6, durante o uso do equipamento e
após sua retirada. O uso do equipamento não causou efeito na produtividade das vacas, nem na qualidade do leite. Algumas mudanças comportamentais foram observadas, mas elas não foram relacionadas a desconforto ou estresse causados pelo uso do equipamento. A realização dos dois primeiros estudos comprova que o melhoramento e manejo de pastagens beneficia o aumento do valor nutricional, o aproveitamento da pastagem no rúmen, com potencial redução de CH4 e promove a
eficiência do comportamento de pastoreio de bovinos. O terceiro estudo valida o uso da técnica SF6 de medição de CH4.
Palavras-chave: Pastoreio. Valor nutricional. Eficiência ruminal. Gases de efeito estufa. Produtividade.
RESUMO EXPANDIDO Introdução
O Brasil é um dos maiores emissores de metano entérico (CH4) no
mundo. O sistema predominante de produção de bovinos no Brasil é extensivo a pasto e caracteriza-se por pastagens degradadas, pouco produtivas e de baixo valor nutricional. Em adição, bovinos em sistemas extensivos têm baixa eficiência de pastoreio e, por serem naturalmente seletivos, promovem excesso de pastoreio, prejudicando o desenvolvimento das plantas e causando degradação dos solos (TEAGUE et al., 2013). Essa realidade pode ser alterada através de estratégias que melhorem a produtividade dos rebanhos e ao mesmo tempo reduzam os impactos ambientais. Sistemas intensivos agroecológicos como o Pastoreio Racional Voisin (PRV) utilizam o melhoramento e manejo de pastagens como estratégia. Como exemplo, cita-se o cultivo de espécies anuais de inverno através da técnica de sobressemeadura, consórcio de gramíneas e leguminosas e controle do acesso do animal ao pasto em seu tempo ótimo de repouso (TOR). Existem diversos métodos para avaliar os resultados da pastagem diante dessas estratégias (HILL et al., 2016). Métodos in vivo são mais
realísticos, mas se desconfia sua influência no comportamento dos bovinos. Métodos in vitro, além de mais acessíveis, mostram alta correlação com as técnicas in vivo (BHATTA et al., 2006). A presente dissertação é composta por três estudos os quais abordam aspectos relacionados a manejo e melhoramento de pastagem em sistema intensivo de PRV, comportamento de bovinos em pastoreio, estratégias de redução na produção de CH4 entérico in vitro e ao uso da técnica in
vivo para mensurar o gás CH4, hexafluoreto de enxofre (SF6).
Objetivos
Os objetivos dessa dissertação foram 1. avaliar o efeito da inclusão de
Vicia sativa L. no cultivo de Avena strigosa Schreb. no valor
nutricional, degradabilidade e produção de gás in vitro da pastagem, e o efeito na seletividade e comportamento de pastoreio de bovinos 2. Avaliar o efeito de diferentes tempos de repouso no valor nutricional, degradabilidade e produção de CH4 in vitro de pastagens polifíticas em sistema de PRV, e o efeito na seletividade e comportamento de pastoreio de bovinos. E 3. avaliar se o uso da técnica SF6 causa
Material e métodos
Os estudos referentes aos objetivos 1 e 2 foram realizados em uma unidade de Sistema PRV da Fazenda Experimental da Universidade Federal de Santa Catarina em Florianópolis com delineamento experimental em blocos completamente casualizados. No primeiro estudo, a área total utilizada foi dividida em seis blocos e cada bloco subdividido em três unidades experimentais de 278 m² cada, nos quais foram distribuídos o cultivo de aveia, cultivo de ervilhaca e cultivo de aveia em consórcio com ervilhaca. A coleta de pasto ocorreu antes da entrada dos animais nas unidades experimentais e foi utilizada para estimar a produção de biomassa e para a realização das análises bromatológicas. Três grupos de quatro vacas eram distribuídos diariamente em cada bloco e unidade experimental em delineamento de quadrado latino duplo 3x3 para pastoreio durante duas horas. Durante esse tempo, as vacas eram observadas e seus comportamentos eram registrados em scans com intervalos de cinco minutos (ALTMANN, 1974). Também foram feitas a simulação de pastoreio de acordo com a técnica descrita por Wallis de Vries (1995), e a contabilização da taxa de bocado cinco vezes por animal por hora durante 30 segundos. Tanto a amostra da pastagem quanto a amostra simulando a seleção dos animais foram submetidas a determinação de matéria seca (MS) e conteúdo de cinzas pela metodologia descrita por AOAC (1995). A determinação de proteína bruta (PB), fibra em detergente neutro (FDN) e fibra em detergente ácido (FDA) foi realizada através do espectrofotômetro infravermelho (NIR). A degradabilidade in vitro e produção de gás da pastagem foram realizadas segundo técnica descrita por Mauricio et al. (1999). A análise estatística dos dados foi feita através do pacote estatístico lme4 (BATES et al., 2015) do R (R CORE TEAM, 2018). Dados da pastagem foram analisados por modelos lineares mistos. Dados de comportamento foram analisados por regressão logística binária (KORNER-NIEVERGELT et al., 2015) multinível (Bernoulli) e a taxa de bocado foi analisada por regressão linear multinível. Valores de P foram obtidos pelo teste tipo II qui-quadrado de Wald (P <0.05 or P <0.01). No segundo estudo foram utilizados 6 piquetes da unidade do PRV como blocos, aos quais foram submetidos 3 diferentes tempos de repouso em subáreas de 834 m² cada: TR1: 24 dias, TR2: 35 dias e TR3: 46 dias. A coleta da pastagem, o estudo de comportamento e as análises de laboratório ocorreram de forma semelhante ao primeiro estudo. Entretanto, no segundo estudo também foi feita a mensuração do gás CH4 por cromatografia gasosa. Análise estatística dos dados foi
semelhante à análise realizada no primeiro estudo. O terceiro estudo foi realizado na unidade experimental da Pontifícia Universidad Católica de Chile, em Pirque no Chile. Vacas leiteiras em estágio de lactação (N=24) foram submetidas ao uso da técnica SF6 (Deighton et al., 2014)
para mensurar produção de CH4 para um experimento à parte. A
produção de leite das vacas foi registrada e amostras foram coletadas para determinar sua composição e estimar a energia corrigida do leite - ECM de acordo com Tyrrell e Reid (1965). O registro da produção e coleta de amostras foram realizados em três diferentes períodos: antes de as vacas serem equipadas com o SF6, durante as mediçõesdo CH4 e após
a remoção do equipamento. A observação individual do comportamento das vacas ocorreu através de gravações por vídeo em quatro fases distintas: antes de as vacas serem equipadas com o SF6 (PRE), enquanto
estavam sendo equipadas (ADAP), durante as medições do CH4
(MEAS) e após a remoção do equipamento (POST). Os comportamentos foram registrados em scans a cada dez minutos. Interações sociais foram registradas durante 1 minuto dentre o mesmo intervalo de tempo. Foi utilizado um datalogger para registrar a frequência com que as vacas se deitavam diariamente. Análise estatística tanto para os dados produtivos quanto para dados comportamentais foi feita através de modelos lineares generalizados (Proc Glimmix) no SAS (SAS institute, 2003). Diferenças entre médias foram avaliadas através do teste Tukey para comparações múltiplas em P <0.05.
Resultados e discussão
No primeiro estudo, a pastagem de ervilhaca e o consórcio de aveia e ervilhaca não diferiram em valor nutricional, mas ambas tiveram maior valor nutricional que a pastagem de aveia; com maior teor de PB (E=24.5 e AE=23.4 vs 14.4 ± 1.36%; P <0.01) e cinzas (E=8.54 e AE=8.73 vs 8.31 ± 0.21%; P <0.01); e menor teor de FDN (E=53.98 e AE=53.40 vs 62.98 ± 1.86%; P <0.01) e FDA (E=31.13 e AE=31.97 vs 34.30 ± 0.62%; P <0.01). O aumento do valor nutritivo da pastagem em razão da inclusão da ervilhaca era esperado, visto que a capacidade de fixação de N atmosférico das leguminosas, naturalmente lhes confere maior teor proteico e menor teor fibroso em sua composição química (MELESSE et al., 2017). No período estudado de 65 dias, a produção de biomassa também foi maior no consórcio e no cultivo de ervilhaca solteira em relação ao cultivo de aveia (1573.12 vs 686 ± 459.5 kg MS/ha; P <0.01, respectivamente). A inclusão da ervilhaca no cultivo de aveia também acarretou melhoria na degradabilidade e parâmetros fermentativos da pastagem. O pastoreio foi o comportamento mais
frequente, sendo que as vacas pastaram mais quando estavam na pastagem composta por aveia e ervilhaca ou só ervilhaca do que quando estavam na pastagem de aveia (93.8, 94.0 e 87.8 ± 0.33%; P <0.05, respectivamente). Entretanto, não houve efeito das diferentes composições de pastagem na seletividade animal e não houve diferença na taxa de bocado (22.3 ± 0.59 bocados/min; P =0.257). Diante disso, assumimos que o aumento na frequência do pastoreio ocorreu em função de os animais terem preferências alimentares, consumindo mais pasto nos tratamentos com ervilhaca. No segundo estudo, houve diferença no valor nutritivo e na quantidade de CH4 produzido. TR1 e TR2 tiveram
teores de proteína (9.51 ± 0.48%) e cinzas (7.61 ± 0.22%) semelhantes, mas TR2 teve maior teor de FDN que T1 (64.56 vs 58.75 ± 1.47%; P <0.01), o que pode ter implicado em maior produção de CH4 (16.33 vs
12.77 ± 0.97 ml/g de MS degradada; P <0.0001). O teor de fibras favorece a atividade de bactérias celulolíticas (VALENTE et al., 2016), as quais produzem maior proporção do acetato, um AGV resultante da fermentação de carboidratos (RIRA et al., 2016). A produção de acetato disponibiliza maior quantidade de hidrogênio (H2), substrato principal
utilizado por bactéricas metanogênicas (KONG et al., 2013). Diante disso, esperava-se que TR3 também mostrasse alta produção de CH4.
Porém, visto que as características fermentativas do alimento estão associadas com seu valor nutritivo, o baixo valor nutritivo de TR3 (PB=8.20, FDN=61.77 e cinzas=6.86) pode ter promovido baixa fermentação e baixa produção de gases. Apesar de não ter sido mensurada a proporção de carboidratos solúveis em cada tratamento, TR3 por ter sido mais longo pode ter tido maior quantidade de fibra indigestível, o que também explicaria a menor produção de gases. A produção de biomassa diária foi maior no TR1 em relação a TR2 e TR3 (44.17 vs 30.40 e 34.43 kg MS/dia). Nos três TR avaliados, o pasto selecionado pelas novilhas teve maior teor de proteína e menor teor de fibra, além de ser mais degradável do que a pastagem ofertada (P <0.01). Isso mostra que nos três TR houve seleção de dieta pelos animais, o que é esperado diante de uma pastagem composta por diversas espécies. Quando há seleção, há aumento na frequência de pastoreio dos animais, como ocorreu no TR3 (66.5 ± 0.11%) em comparação a TR1 (63.5 ± 0.08%) e TR2 (64.3 ± 0.10%) (P <0.05). Também notamos diferença na taxa de bocados, a qual foi maior em TR1 em comparação a TR2 e TR3 22.18 (±0.60); 18.40 (±0.71) e 18.65 (±0.71). No TR3, em função do pasto ter baixo valor nutricional e maior disponibilidade total de biomassa, as vacas tiveram que selecionar mais o pasto colhido, e para compensar a ingestão, se dedicaram mais
tempo na busca de seus alimentos (CORNELISSEN & THEO 2015). No terceiro estudo, não houve diferença na produção de leite (32.2 ± 1.6 kg/d) nem em sua composição, o que nos permite assumir que não houve alteração na produtividade das vacas causada por estresse relacionado ao uso do equipamento SF6 (RHOADS et al., 2009;
BROUCEK et al., 2017). Em relação ao comportamento, não houve diferença no tempo total de descanso das vacas (12.07 ± 0.35 h/d;), apenas uma tendência de redução durante a fase ADAP (P =0.06). Essa tendência pode ter sido apenas uma resistência das vacas no momento de introdução ao equipamento, por ser algo incomum de sua rotina. As vacas ruminaram mais durante a fase MEAS (15.1 vs. 12.1 ± 0.64 min/h; <.0001) e não houve diferença na frequência de alimentação das vacas. Não houve diferença na frequência de interações agonísticas entre as fases. Entretanto, as interações afiliativas aumentaram (P =0.02) durante a fase ADAP (1.65 nº/h) e reduziram durante a fase MEAS (0.85 nº/h). As vacas tendem a se aproximar de suas vizinhas como suporte social frente a situações que lhes parecem desafiadoras (RAULT, 2012) Considerando que o equipamento era uma novidade, elas podem ter procurado esse suporte durante a fase ADAP.
Considerações finais
As estratégias de melhoramento e manejo de pastagens testadas nessa dissertação foram eficientes para melhorar o perfil nutricional do alimento e os parâmetros fermentativos do rúmen, com potencial redução do CH4 in vitro. Essas estratégias afetaram o comportamento de
pastoreio dos bovinos ressaltando suas preferências alimentares através de mudanças na frequência do pastoreio. A ação da seletividade dos bovinos também foi destacada. A hipótese do terceiro estudo era que o uso da técnica SF6, validada para mensuração do CH4, iria alterar a
produtividade e o comportamento de vacas leiteiras. Essa hipótese não foi confirmada. As pequenas alterações comportamentais observadas ocorreram apenas na fase de adaptação das vacas em relação ao equipamento. As estratégias de pastagem avaliadas nessa dissertação apresentaram potencialidade de redução do gás CH4 de bovinos em
sistemas intensivos de pastoreio, e a técnica in vivo de mensuração de gases foi validada por não mostrar ter causado estresse às vacas durante a fase de medições do gás e nem redução na produção de leite no período avaliado.
Palavras-chave: Pastoreio. Valor nutricional. Eficiência ruminal. Gases de efeito estufa. Produtividade.
RESUMEN
El mejoramiento de praderas incrementa el valor nutricional de forrajeras, mejora la degradabilidad en el rumen y la eficiencia del comportamiento de pastoreo. El cultivo de especies anuales de invierno en asociación proporciona beneficios a los animales y al ambiente. El mejoramiento debe ser acompañado por un manejo adecuado, respetando la dinámica de las praderas, permitiendo que sean pastoreados por los animales en su tiempo óptimo de reposo, lo cual implica en una pradera de mejor calidad, como también influencia en la producción de metano entérico ruminal (CH4). Formas in vitro de
estimar la producción de CH4 se basan en la simulación de fermentación
ruminal y formas in vivo lo hacen directamente del animal. Todavía no se sabe la influencia de esas técnicas in vivo en el comportamiento de los animales. Esta investigación es compuesta por tres estudios. El primer objetivo fue evaluar el efecto de inclusión de ervilhaca (Vicia sativa L.) en el cultivo de avena (Avena strigosa Schreb.), en el valor nutritivo y degradabilidad in vitro de la pradera y su efecto en el comportamiento de novillas. Por tanto, se realizó el cultivo de avena sola, avena en asociación con ervilhaca y ervilhaca sola en un área dividida en 6 bloques, siendo colectadas las muestras de pasto para los análisis bromatológicos y de degradabilidad in vitro. Fue observado el comportamiento de novillas durante dos horas y realizada la simulación de pastoreo y la contabilidad de bocados. La inclusión de leguminosas aumento el tenor de proteína de la avena y disminuyo los tenores de fibra, promoviendo mejor degradabilidad in vitro, además de aumentar la eficiencia de pastoreo. El objetivo del segundo estudio fue evaluar el efecto del manejo en el valor nutricional de la pradera, degradabilidad y producción de CH4 in vitro y la influencia en el comportamiento de las
novillas. Para ello se evaluó tres tiempos de reposo: TR1: 24 días, TR2: 35 días y TR3: 46 días, siendo realizadas colectas de pasto para análisis bromatológico, degradabilidad y producción de CH4 in vitro. Las
observaciones de comportamiento se realizaron durante cuatro horas, acompañadas de simulaciones de pastoreo y contabilidad de bocado. El TR1 se destacó por tener más proteína, menos fibra y menor producción de CH4. Hubo efecto de selectividad animal en la calidad de forrajeras y
en la degradabilidad, pero eso no se encontró en la producción de CH4.
La mayor taza de bocados ocurrió en el TR1, pero la frecuencia de pastoreo fue mayor en el tiempo más tardío. El tercer estudio tuvo como objetivo evaluar el efecto de la técnica de hexafluoreto de azufre (SF6)
productividad y comportamiento de vacas lecheras. La producción de la leche se registró y se tomó muestras para el análisis de calidad. El comportamiento de las vacas fue evaluado y comparado antes del uso del equipamiento, mientras que ellas estaban siendo equipadas con el SF6, durante el uso del equipamiento y después de la retirada. El
equipamiento no tuvo efecto en la productividad de las vacas, ni en la calidad de la leche. Fueron detectados pequeños cambios comportamentales, los cuales no fueron asociado con estrés por el uso del equipamiento SF6. La realización de los dos primeros estudios
comprueba que el mejoramiento y manejo de praderas mejora la calidad y aprovechamiento ruminal de las forrajeras, con potencial de reducción de CH4, así como promueve la eficiencia del comportamiento de
bovinos. El tercer estudio valida el uso de la técnica SF6 de medición de
CH4.
Palabras-clave: Pastoreo. Valor nutritivo. Eficiencia ruminal. Gases de efecto invernadero. Productividad.
ABSTRACT
Pasture improvement is a strategy to increase the pasture nutritional value that will improve the degradability on rumen and maximize the grazing efficiency. One of the more popular strategies recommended is that of cultivating two winter species in one pasture. This provides not only benefits to the cattle, but also provides a healthier and sustainable pasture. However, adequate management that respects the pasture growth dynamics is needed. That must allow for grazing only after reaching the optimum recovery period. The optimum recovery period not only implies a better quality pasture, but also influences in the production of enteric methane (CH4). In vitro techniques of measuring
CH4 production are based on the simulation of ruminal fermentation,
whereas in vivo techniques rely on the direct emissions from the animal. However, the influence of techniques that test the natural emission of CH4 on the animal’s behaviour is unknown. This thesis is composed of
three studies. The first study is aimed at evaluating the effect of the combining of common vetch (Vicia sativa L.) with black oat cultivation (Avena strigosa Schreb.), regarding nutritional value and in vitro degradability of the pasture, as well as the effects on heifer behaviors. Considering this, oats grown alone, oats grown with vetch, and vetch grown alone were used in an area divided into six even blocks. In these six blocks they were divided into thirds, and each received different distributions of the above three categories of grasses. Pasture samples were collected for bromatological analysis and in vitro degradability. The behaviour of the heifers was observed for two hours, while grazing, hand-plucking, and bite rates were measured. The inclusion of common vetch significantly increased the oat protein content and decreased the fiber content, which promoted better in vitro degradability and increased the grazing efficiency. The second study is aimed at evaluating the growth recovery rate of naturally grown pastures, grass quality of the pastures, degradability, CH4 in vitro production, and the heifers'
behaviour. Due to this criterion, three recovery periods were evaluated in six different equal blocks on a rotation of 24 days, 35 days and 47 days. We analyzed pasture samples, degradability, and the CH4
production in vitro. We also observed and measured the behaviour of the cows for four hours, using hand-plucking and the bite ratio. During the 24-day period, we noted having better nutritional grass with more protein and less fiber content and a lower production of CH4. Out of the
three selected groups of six cows per group all selecting their preferential pasture samples with different qualities of grass this did not
change the emission of CH4 production in any of the pasture samples.
Higher bite rate occurred during the earlier recovery period, but the frequency of grazing was higher during the later recovery period. The objective of the third study was to evaluate the effect of the in vivo sulfur hexafluoride (SF6) technique to measure CH4 production on dairy
cows' productivity and behaviour. Milk production was registered, and samples were taken to quantify the milk solids yield and quality. The cows' behaviours were evaluated and compared before using the SF6
equipment, then while they were fitted with SF6, equipment, during the
use of the equipment, and after the equipment removal. The use of the equipment did not affect milk yield and quality. Some minor behavioral changes were noticed, which did not negatively impact the cows, showing that they were able to adapt to the SF6 equipment. The
conclusion of the first two studies proves that the improvement and careful management of pastures improves the quality and the ruminal efficiency of grazing, with the potential reduction of CH4, as well as the
promotion in the efficiency of cattle behaviour. The third study validates the use of the SF6 technique to measure CH4.
Keywords: Grazing. Nutritional value. Rumen efficiency. Greenhouse gases. Performance.
LISTA DE FIGURAS
Figure 1. The gas volume (A) and DM degradability (B) in each grass category according to incubation time…...………60 Figure 2. Pasture degradability (%) according the incubation time (6, 24 and 48 hours) and the sampling pasture by square method and by the hand-plucking method. P <0.05.………...…………...73 Figure 3. Cow equipped with the Sulphur hexafluoride (SF6) equipment
to measure enteric methane (CH4) emissions. From left to right: the
saddle with PVC canisters, leather head halter with the sampling point, and the entire equipment.………...81
LISTA DE TABELAS
Table 1. Response of variables obtained through the linear mixed-effects models applications in relation to: O – oat grown alone, O+V – oat grown with vetch and V – vetch grown alone, and to the cow’s selection: Square pasture samples from paddocks or hand-plucking (HP) samples selected by cows. CP: Crude protein (%), ADF: Acid detergent fiber (%), NDF: neuter detergent fiber (%), Ashes (%), and biomass production (DM/ha). N=36……….59 Table 2. Response of variables obtained through the linear mixed-effects models applications in relation to: RP1 – 24 days, RP2 – 35 days, and RP3 – 46 days, and to the cow’s selection: Square pasture samples from paddocks or hand-plucking (HP) samples selected by cows. CP: Crude protein (%), ADF: Acid detergent fiber (%), NDF: neuter detergent fiber (%), Ashes (%), and biomass production per day (DM/ha/day). N=36………...72 Table 3. Production of methane (CH4) gas (ml) per DM degraded (g) in
relation to: RP1 – 24 days, RP2 – 35 days, and RP3 – 46 days and according to the incubation time (6, 24 and 48 hours). Different letters mean significance difference………..…73 Table 4. Cow production and behavior in each of four phases of fitting with sulfur hexafluoride (SF6) equipment. PRE: Before cows had the
equipment on; ADAP: initial period of fitting; MEAS: During the enteric methane (CH4) measurements; POST: after the SF6 equipment
LISTA DE ABREVIATURAS E SIGLAS ADAP While cows were fitted with the SF6 equipment
ADF Acid detergent fiber C Carbon
CEUA Ethic Committee of Animal use CH4 Enteric methane
CO2 Carbon dioxide
CP Crude protein DM Dry matter
ECM Energy corrected milk GHG Greenhouse gas H2 Hydrogen
LETA Applied ethology laboratory
LQAP Analytical Chemistry of Plants Laboratory MEAS During the CH4 measurements
MSY Milk solids yield MY Milk yield N Nitrogen
NDF Neutral detergent fiber NH3 Ammonia concentration
NIR Near infrared spectrometry ORP Optimum recovery period
POST After the SF6 equipment was removed
PRE Before cows were fitted with the SF6 equipment
RP Recovery period
RP1 Recovery period of 24 days RP2 Recovery period of 35 days RP3 Recovery period of 47 days SF6 Sulfur hexafluoride
TDN Total Digestive Nutrient
UFSC Federal University of Santa Catarina UNB University of Brasilia
VFA Volatile fatty acids VRG Voisin’s Racional grazing
SUMÁRIO 1INTRODUCTION ... 37 2OBJECTIVES... 41 2.1General objective ... 41 2.2Specific objectives ... 41 3LITERATURE REVIEW ... 43 3.1Pasture improvement and management ... 43 3.2 Grazing behavior and pasture selectivity ... 45 3.3 Ruminal fermentation ... 47 3.4 Enteric CH4 estimation and measurement on ruminants ... 49 3.5 Influences that factor in dairy cow’s behavioral changes in research………….. ... 51 4 BLACK OATS GROWTH WITH THE COMMON VETCH TO IMPROVE THE IN VITRO DEGRADABILITY AND GAS PRODUCTION OF PASTURE ... 53 4.1Introduction ... 53 4.2Material and methods ... 54
4.2.1 Site description ... 54
4.2.2 Animal, treatment and experimental design ... 55
4.2.3 Collection of pasture and samples ... 55
4.2.4 Behavioral observations and hand-plucking ... 56
4.2.5 In vitro gas production technique ... 56 4.2.6 Statistical analysis ... 57
4.3 Results ... 58 4.4 Discussion ... 60 4.5 Conclusion ... 63 5 EFFECT OF RECOVERY PERIOD ON IN VITRO METHANE EMISSION FROM MIXTURE PASTURE ... 65 5.1Introduction ... 65 5.2Material and methods ... 67
5.2.2 Animals and experimental design ... 67 5.2.3 Pasture collect and samples... 68
5.2.4 Behavioral observations and hand-plucking ... 68
5.2.5 In vitro gas production technique ... 69 5.2.6 CH4 measurements ... 70
5.2.7 Statistical analysis... 71
5.3Results ... 71 5.4 Discussion... 73 5.5Conclusion ... 77 6 MILK PRODUCTION AND ANIMAL BEHAVIOR ARE NOT AFFECTED BY THE EQUIPMENT USED IN THE SF6 TECHNIQUE TO ESTIMATE METHANE EMISSIONS FROM DAIRY COWS ... 79 6.1Introduction ... 79 6.2Material and methods ... 80
6.2.1 Site description ... 80 6.2.2 Animals and experimental design ... 80 6.2.3 Milk samples ... 81 6.2.4 Behavioral evaluations ... 81 6.2.5 Statistical analysis... 82 6.3Results ... 82 6.4Discussion ... 83 6.5Conclusion ... 85 7GENERAL DISCUSSION ... 87 8REFERENCES ... 91
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1 INTRODUCTION
Brazilian agriculture is very important, and it contributes actively to our economic growth (IBGE, 2017). Cattle are ranked at the top of the Brazilian agriculture market and we are one of the world’s largest suppliers of beef (FAOSTAT, 2017). Cattle are raised mainly in pasture systems, predominantly extensive ones, occupying vast areas of Brazilian lands (PEREIRA et al., 2018). However, due to the inadequate handling and control of these animals and pastures, Brazil’s systems are inefficient because of low nutritional and poorly managed pastures (BERNDT; TOMKINS, 2013; MILLEN; ARRIGONI, 2013). In addition to this, pastures are mainly composed of native species of grasses that have low productivity and offer instability during the year (FERRAZ et al., 2010). These circumstances create a low efficiency for the production of Brazilian beef, which despite how popular Brazilian beef is worldwide, we need to figure it out a way to create higher productivity (SALTON et al., 2014).
In extensive systems the animals have the possibility to walk around, choose, and select the area in which they are going to graze (DEVRIES; DALEBOUDT, 1994). Then they will probably come across a high fiber and low nutritional pasture, this will decrease efficiency because they will spend more time selecting proper grazing area rather than grazing, thus compromising the herd productivity. Moreover, since cattle are very selective and have food preferences (ANIMUT et al., 2005), they end up overgrazing in main areas. They prevent plant recovery, prejudicing their radicular development and causing soil degradation (TEAGUE et al., 2013), making the system even less productive. For this reason, production systems have been redrawn in order to find more sustainable strategies that improve their efficiency (SALTON et al., 2014), and respect ecosystem dynamics.
An efficient strategy that is easily available and cheap is simply pasture improvement, which promotes healthier more resilient and better nutritional pastures (TEAGUE et al., 2013). In agroecological intensive systems, this strategy is a premise to achieve high productivity and, at the same time, reducing environmental impact by promoting more sustainable systems (NIGGLI et al., 2009).
Well managed pastures have a higher soil carbon (C) sequestration through their roots (CHEN et al., 2015); this improves the radicular development and provides a nutrient cycle and a biodiversity establishment (NIGGLI et al., 2009). They also provide an organic matter accumulation for the soil (TEAGUE et al., 2013), which
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stimulates the microorganismal process for creating soil biocenosis (NIGGLI et al., 2009).
Improved grazing regimes have vigorous growth, providing cattle with a better nutritional diet, thus influencing their behavior and providing grazing efficiency (MCCARTHY et al., 2007). Better nutritional diets stimulate animal hunger and improve feed efficiency, are more digestible in the rumen, and create better fermentation parameters with maximum nutrient absorption (WANAPAT, 2000). Thus, to improve animal productivity, to minimize their enteric methane (CH4) production (KNAPP et al., 2014), which represents between 2%
and 12% of the gross energy intake of the animal (JOHNSON; JOHNSON, 1995).
CH4 is a greenhouse gas (GHG) that ferments in the rumen and
becomes one of the most important problems in raising cattle. Considering that Brazil, plays a big role in worldwide beef production, it is positioned among the biggest CH4 emitters in the world (FAOSTAT,
2016). Therefore, researchers are looking for strategies to evaluating and comparing nutritional values and digestibility from different grasses and legume species, as well as, the animal’s acceptance and the digestibility in the rumen, thus reducing the emissions. There are many methods to accomplish this, and some still offer more detailed results from ruminal fermentation, such as gas production and enteric CH4
concentration (HILL et al., 2016).
Methods that measure CH4 directed from animals is more
realistic. Nevertheless, their use might be unfeasible due to the high numbers of animal demand of cost and time. Furthermore, it is not known if these methods affect the animal’s behavior, which somehow could generate erroneous results. As an alternative, there are in vitro methods that are more accessible and respect the 3Rs principles
(replace, reduce, refine) in animal research (BOO et al., 2005). In vitro
techniques simulate the ruminal process in the laboratory based upon the evaluation of different substrates and feed regarding fermentation, degradability, metabolized energy, and gas production (PALIC; LEEUW, 2009). These techniques have the effects that might change the fermentation condition, such as the pH and rumen, and the liquid composition, under control (STORM et al., 2012), and show high correlation with direct techniques (BHATTA et al., 2006).
The present thesis is aimed at evaluating aspects related to pasture management in Voisin’s Racional Grazing (VRG) system. This aspect is found to improve the pasture nutritional value, degradability, and grazing efficiency, including the management potentiality in
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reduced enteric CH4 in vitro production. Additionally, we aimed at
evaluating some behavioral aspects of the grazing habits of our test cattle, their capability to select their pastures, to ingest, and the impact of the sulfur hexafluoride (SF6) technique used to measure the CH4 on
the dairy cow’s behavior. Therefore, this thesis is composed of five main textual chapters. The first – the review of main topics from journals that support this project: pasture improvement and management, cattle grazing behavior, ruminal fermentation, techniques of enteric CH4
measurement, and the estimation of factors that might have influence on dairy cows' behavior that are submitted before research. Chapters four through six approach the studies that developed the composition of this thesis. Chapter four is about the cultivation of mixed pasture species in the VRG system and its impact on pasture nutritional value, degradability, and on grazing efficiency. Afterwards, chapter five is about the importance of pasture management dealing with the CH4
production and emissions. Chapter six shows us the changes in the dairy cattle's behavior when they are submitted to SF6 technique. Finally,
chapter seven shows us a summary discussing all the previous chapters and their final conclusions.
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2 OBJECTIVES 2.1 General objective
The evaluation of the pasture strategy management with regards to the in vitro fermentation and pasture degradability, such as the impact on cattle grazing. Additionally, to measure the SF6 technique and check
if it is a viable measurement that does not change the cow’s productivity, and their usual behavior.
2.2 Specific objectives
1) Evaluating the effect of common vetch on the oat growth regarding degradability and gas production, as to how it affects cow selectivity and grazing behavior;
2) Evaluating the effect of three recovery periods of mixed pasture regarding chemical compositions, in vitro CH4 production and
degradability, as to how it affects cow selectivity and grazing behavior; 3) Evaluating if the use of the SF6 technique implies that dairy
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3 LITERATURE REVIEW
3.1 Pasture improvement and management
Agriculture is a very important worldwide activity occupying vast areas of land throughout the world (FAOSTAT, 2016). Agricultural systems must be agroecologically intensified towards profitability and the increase in resilience (CONANT; PAUSTIAN; ELLIOTT, 2001), in a manner that reduces all the negative impacts such as water pollution, soil degradation, deforestation, and GHG emissions coming from certain activities (NIGGLI et al., 2009). Grass-based systems need to be extremely high in nutrients so that the cattle will produce well. Once a pasture operates to its optimal performance, then it is required the pasture to be well managed and offer a high quality and productive diet to the cattle. As a result, broad pasture management will bring benefits to the animals, the farm, and the environment (GREGORINI et al., 2017).
Pasture improvement practices include the choice and the variety of productive high valued nutritional species that suits the environmental characteristics, the use of plants that cover the soil; the use of legumes that generates the highest concentration of nitrogen (N) on soil, crop rotation (WOODWARD et al., 2001); and the combination of grasses that can grow together without harming one another, under no-tillage soil (ORDÓÑEZ et al., 2018).
Sowing a winter species is a common practice that increases nutritional value and productivity in pastures during the winter, when native tropical pastures have unstable winter yields (CHAMAILLÉ-JAMMES; BOND, 2010). Winter species are usually seeded over the natural pasture (MOREIRA et al., 2005), which keeps the native vegetation and soil intact and avoids soil erosion (HEINRICHS et al., 2001), as well as benefiting exchange of nutrients for the plant roots. To maximize this strategy, the mixture of species and legume inclusion is recommended.
Plant species have distinct radicular systems. When working in cooperation with other species, this stimulates each plant's process and creates the development of soil microorganisms (NIGGLI et al., 2009). Therefore, a nutrient cycle (NIGGLI et al., 2009), and organic matter soil accumulation (TEAGUE et al., 2013) will be guaranteed, providing soil fertility, better establishment of different species in pastures, and avoiding the loss of N through soil lixiviation (TILMAN et al., 1996). Additionally, mixed species are more resistant to weed invasion
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(STURLUDÓTTIR et al., 2013), and promotes a better N and fermentable energy intake balance (PEYRAUD; DELAGARDE, 2011). Pasture nutritional value is increased with legumes due to their higher protein content compared to grasses, improving the C/N rate and herbage digestibility. The increment of N on the soil improves its fertility and the nutrient availability to plants, increasing pasture productivity, which also reduces the need for chemical fertilizers (ANJOS et al., 2016). Some legumes are also full of condensed tannins (WOODWARD et al., 2001) and secondary metabolites, that change the N metabolism and the ruminal microflora, also reducing the enteric CH4
production (BROUDISCOU; PAPON; BROUDISCOU, 2000).
Black oats (Avena strigosa Schreb.) are commonly considered a winter grass used in Southern Brazil, as is the common vetch (Vicia
sativa L.), a legume. The oats provide increased productivity and slower
pasture decomposition under the soil, protecting it against erosion. Common vetch improves pasture's nutritional value and the N availability for the future plant growth, due to its ability N fixation from the atmosphere (HEINRICHS et al., 2001).
Mixing grasses and legumes also contributes to the organic matter soil conservation which, through photosynthesis, creates an ability to saturate C into the soil. This compensates for the historical losses caused by deforestation, overgrazing, and inadequate handling of pastures throughout history (CONANT; PAUSTIAN; ELLIOTT, 2001). When the accumulated C in the soil is higher than the amount emitted from the ruminant’s production, there is a negative balance regarding GHG emissions (CHEN et al., 2015). Likewise, this accumulation of C in the soil must overcome the losses of C through soil oxidation and erosion (SILVA et al., 2016).
Pasture improvement will bring plenty of benefits to the animals and the longevity and productivity of the pasture system. Nevertheless, these benefits require the pasture to be handled correctly and frequently. Handling consists of planning biomass availability and controlling access to the pasture. Rotational systems, like the VRG system, or the Voisin system (FARLEY et al., 2012; MACHADO FILHO, 2011; TEAGUE et al., 2013), provides this management instruction. The VRG system is divided into several paddocks, where the cattle access is rotated on a daily basis so that they have a new pasture available every day. Therefore, the cattle's access to this paddock pastures will always be at the optimum recovery period (ORP) of the pasture. The pastures at ORP produce higher productivity, higher nutrient content and increase digestible matter proportion of the plant
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per time and per area (MACHADO FILHO, 2011). Using the ORP formula allows for leaving animals in the paddock for a shorter period of time with higher stocking rate, which also provides time for the pasture to recover on a regular time schedule (TEAGUE; DOWHOWER, 2003).
Recovery periods (RP) in extensive systems are very tricky. The pasture is not managed and has low productivity and low nutritional value (VOISIN, 1961). Moreover, since the animals have a food preference and are naturally capable of selecting their pastures (ANIMUT et al., 2005), they usually choose plants that are more appetizing and have the newest leaves. This promotes overgrazing and soil degradation (TEAGUE et al., 2013). When the RP is late, plants start coming into their reproductive cycle, directing their nutrients to the flowering and seed formations (MACHADO FILHO, 2011). The protein content is expected to decrease at the same time that the cell wall content is expected to increase, making the pasture have a higher content of fiber. A high-fiber content pasture promotes the pH increment of the rumen and the rumen retention time of the pasture will also increase. This decreases the animal feed efficiency and changes the fermentation end products (BEAUCHEMIN et al., 2008). Hence, the CH4 production
is higher compared to pastures that provide a higher protein content and easier digestibility (JOHNSON; JOHNSON, 1995).
3.2 Grazing behavior and pasture selectivity
Grazing behavior is the most expressive behavior in cattle when they are raised on grass-based systems (ANDRIAMASINORO et al., 2016). Grazing behavior encompasses the search, the selection, the intake, and the satiety of food, which leads to the ruminating process. These activities result from an interaction between the cattle, the pasture, and environmental factors (BARRETT et al., 2001), and are aimed at balancing animal nutritional levels, animal energy demand, and animal satiety (GREGORINI et al., 2017).
Integrating the interaction of factors that make grazing efficient, we need to consider the cattle, the environmental elements, and the plant species. Cattle choose where they graze based upon their nutritional needs at any specific time (GREGORINI et al., 2015). The choice of the animal depends upon the different grass distributions in the pasture, the plant height, the presence of secondary metabolites (MEISSER et al., 2014), and the plants ability for photosynthesis (MANNING et al., 2017). Cattle select different parts of plants, dependant upon whether the
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superior or inferior parts are preferred for nutritional value (BENVENUTTI et al., 2015). Some variables besides grazing frequency, such as rate and volume of bite and rate of intake, provide us with observations about the animal selection when grazing. These variables are controlled by physical intrinsic signs in the animal, such as their requirement for nutrients and the ruminal filling, and in these situations cattle generally become more selective according to their satisfaction that specifically relates to what they have eaten (GREGORINI et al., 2007). Intensive systems like VRG predict paddock changes daily, and the cattle graze in a different pasture daily; cattle will eat voraciously in a short period of time, trying to compensate for the paddocks change, thus minimizing or avoiding their natural ability to select different species of plants (MACHADO, 2010).
The importance of the hierarchy in the herd influences when, where, and what time the cow eats. Dominant cows have priority when accessing resources. This means that they have the liberty of to choose the best areas which they deem beneficial (PHILIPS; RIND, 2002). Whereas, submissive cows are forced to graze in areas left over or have to wait until the dominant cows are finished grazing in particular areas (SOWELL; MOSLEY; BOWMAN, 2000). When time is limited, as is common in intensive systems, the dominant cows harm the behaviour of the submissive cows thus reducing their bite rate and diminishing their grazing time (SOWELL; MOSLEY; BOWMAN, 2000).
Cattle behavior is also molded according to the plant species and their characteristics in a particular pasture (BENVENUTTI et al., 2015). This is judged upon the leaf-stem rate, the biomass production, and the phenological stage (ORR et al., 2004). Botanical plant characteristics are influenced by the seasonality of the year (ENRIQUEZ-HIDALGO et al., 2014) and by the management they receive. This impacts the quality of the animal’s intake and influences the ruminating rate. The ruminating function is to reduced to the food particle size and facilitates the degradation, the digestion, and the nutrient absorption. This will influence the animal’s nutritional efficiency and the volatile fatty acids (VFA) production (MENDES et al., 2014). The average cow spends 33% of her time ruminating (KUMAR; HANCKE, 2015). This time can be altered due to different feed factors, such as fiber content, lignin, intake, and stressful factors (AMBRIZ-VILCHIS et al., 2015). Ruminating is also associated with the cow’s ability in resting (KUMAR; HANCKE, 2015). Cattle prefer to ruminate in a restful position, such as lying down (SCHIRMANN et al.,
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2012). When any of these behaviors are changed, it can indicate discomfort or welfare issues.
The biomass availability is also a very important influential factor upon grazing behaviors and must agree with the rate of stocking cattle and the time they will graze in each area. The biomass will depend upon the plant species, the time of year, and where they are grazed. When the pasture offers high biomass, animals tend to have brief meals, with a higher volume of bite and less bite rate (MOTUPALLI et al., 2014) whereas, when there is low biomass, the animal needs to search and walk more, expending more energy, which also requires a higher bite rate and a lower volume of intake in each bite (MEZZALIRA et al., 2014). Time dedicated to grazing also tends to be higher when the biomass availability is low. However, this does not imply higher foraging, since searching for food is accounted as part of their natural grazing behavior.
Grazing behavior is an innate behavior of cattle and represents a huge part of its daily activity. When cattle are not grazing or ruminating, they are dedicated to other activities like (MENDES et al., 2014), the search of water, shade, places to rest, and social interactions. It is extremely important to comprehend the factors that influence the cows' expressions. All these activities have an impact on the animal’s performance (BENVENUTTI et al., 2015).
3.3 Ruminal fermentation
Rumen fermentation is a part of the natural digestion in cattle. The main function is fermentation of carbohydrates typically, VFA production, amino acids synthesis from non protein N compounds, B complex synthesis, and synthesis of vitamin K (VALENTE et al., 2016). Rumen in its normal condition has a pH between 6 and 7, low oxygen concentration, and a temperature between 38 and 42°C (HILL et al., 2016). These conditions must be kept during food intake, in order to guarantee the efficiency of the digestion process and favor the survival of the beneficial microorganisms.
Rumen efficiency is expected to increase the pH after fermentation, which betters fiber degradation, promotes higher VFA production, higher digestibility of nutrients, N retention, and a higher concentration of ammonia (NH3) (MOHAMED et al., 2009). N and NH3
are substrates used by bacteria to create protein, which stimulates growth and provides a higher protein quality (GIANG et al., 2016). The concentration of N and NH3 comes from protein metabolism and is a
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source of cellulolytic bacterial growth. This bacterium is responsible for the degradation of cellulose from the plant cell wall, and their survival is affected when there is a higher concentrated amount of feed included in the diet, promoting the break down of the pH and fermentation more quickly (VALENTE et al., 2016). The prolonged fall of the pH will harm the microbial metabolism, the DM intake, and food degradability, and will also cause illnesses, such as acidosis, inflammation, and diarrhea (CHAUCHEYRAS-DURAND; FONTY, 2002).
Cellulose degradation is a part of carbohydrate fermentation, which results in VFA production - the main energy source for ruminants, and substrates to the metabolic paths (DEPPENMEIER, 2002). The main VFA are produced as acetate, propionate, butyrate, isobutirate, isovaleric, and formate; but acetate and propionate are produced in bigger proportions (MOSS; JOUANY; NEWBOLD, 2000). VFA is also used in other metabolic paths. Some of them act as an energy source in the methanogenic pathway, which is where CH4 is produced (KONG et
al., 2013). This occurs in anaerobic condition 22because of the Archaea
bacteria from the Methanobrevibacter or the Methanosphaera genus (POULSEN et al., 2013). Ciliate protozoa and anaerobic fungus also participate, but in lower proportions (TAPIO et al., 2017). Due to the high bacterial specificity, different compost can be used to produce CH4.
Most bacteria use hydrogen (H2) electrons to reduce the carbon dioxide
(CO2) in CH4, but sometimes CO2 electrons oxidize reducing the methyl
groups to methylamine and methanol to CH4 (JANSSEN; KIRS, 2008).
The VFA acetate butyrate can also be converted into CO2 and CH4, but
propionate is our goal to reduce the CH4 gas. For propionate to be
produced available H2 is needed, thus producing more propionate, less
H2 is available for the methanogen bacteria (MOSS; JOUANY;
NEWBOLD, 2000), resulting in less CH4 emissions.
The profile of VFA produced and the ruminal microbial dynamics are usually influenced by the type, the quality, and the amount of food intake of the animal (O’MARA, 2011), which dictate the time of the digested process (MOSS; JOUANY; NEWBOLD, 2000). Considering that the H2 is the main substrate used by bacteria to produce
CH4, which increase the opportunity of the alternative metabolic path
for the H2, is a strategy to mitigate enteric CH4 (POULSEN et al., 2013).
This includes cattle fed with food that promotes higher production of propionate over acetate (WANG et al., 2016).
Different plant species have different chemical profiles; we differentiate according to digestibility and fermentation rate (BUDDLE et al., 2011). Therefore, mixing many varied species promotes higher
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nutritional value in pastures, and also increases the benefits upon the environment (THORNTON; HERRERO, 2010). Overall, the C3 carbon pathway species, like the temperate climate plants (PRIMAVESI et al., 2004) and legumes (WOODWARD et al., 2001) tend to generate less CH4 production. Therefore, the choice of species and pasture
management are very important in grass-based systems. Additionally, adequate pasture management contributes to the best nutritional value of the pasture and could have an impact on lower CH4 emissions per kg of
dry matter (DM) intake.
3.4 Enteric CH4 estimation and measurement on ruminants
There are several methods to measure and estimate enteric CH4
production on ruminants. These methods are based in equipment, laboratory techniques, tracers, sensors and mathematical models (HILL et al., 2016). They also have a distinct advantage and disadvantage, compliant with regards to different conditions and restrictions that are typically used.
The in vitro techniques are economically accessible and easily executed. They have a low co-efficient of variation since they are in a controlled environment. These techniques do not demand more than two or three animals to collect their ruminal fluid. In vitro techniques simulate rumen fermentation and allow for the inference and feed degradability, all the while considering gas production (PALIC; LEEUW, 2009). This enables the use of several foods and substrates in different proportions and combinations, beside offering data and information about kinetic fermentation (MAURÍCIO et al., 2003). There is a high correlation between in vitro and in vivo techniques (BHATTA et al., 2006); that is why pre-evaluation in the laboratory is recommended (DOREAU et al., 2016), before we do in vivo.
According to the in vivo method, the metabolic chambers, or also called the fermentation chambers, and are used regularly. These chambers work as open respiratory circuits that detect all the heat and the gas production, including the CH4; and they also detect the complete
animal energy metabolism. (STORM et al., 2012). It is also possible to collect the urine and the feces from the animal, thus necessitating the estimation of the digestibility of the feed that has been consumed (HILL et al., 2016). Likewise, animals kept in a closed environment, are observed according to daily ingestion, weight gain, and individual productive characteristics (PIÑARES-PATINO et al., 2011).
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The metabolic chamber results are commonly used in a developed mathematical model, and these equations estimate ruminant emissions This data is compilated and used in the GHG national inventory (HILL et al., 2016). The chamber fermentation method has been criticized for restricting the typical cattle behavior, simply because the argument has been based upon an artificial environment, which is divergent from the natural environment where cattle are typically raised (PIÑARES-PATINO et al., 2011). Restricting pattern behaviors can generate erroneous data with a high co-efficient of variation (STORM et al., 2012). Therefore, in order to use this method in cattle without needing to lock them up or cloister them, the SF6 tracer method was
developed to measure the CH4 emissions (JOHNSON et al., 1994). The
SF6 method of tracing, uses SF6 inert gas which includes similar
characteristics such as CH4, and estimates the concentration of CH4
emitted by each animal in what we call chromatography (JOHNSON et al., 1994). A permeation tube is inserted into the rumen and loaded with SF6, which releases a known rate of gas (JOHNSON; JOHNSON, 1995).
The release of the SF6 into the rumen allows the captured sample of the
dissipated atmospheric dilution observed around the mouth and the nose. The collection of a sample can be collected through a capillary tube, the size and thickness of which, can influence the sample of the gas flow (JOHNSON; JOHNSON, 1995). The capillary tube is usually fixed to a halter on the animal’s head and is connected to an airtight cannister (JOHNSON; JOHNSON,1995). The canisters are made of PVC, and originally were put around the animal’s neck (HILL et al., 2016). However, there are adaptations of techniques in order to make the equipment more comfortable for the animal. The technique used by Deighton et al. (2014), for instance, put the canister mounted on a padded flexible accessory that is fitted to the back of the cow. There is a plastic strap that is an accessory that is placed around the cow’s hindquarters. Samples can also be collected through an orifice plate instead of a capillary tube.
The SF6 tracer method is efficient to measure the CH4
production, and this method still generates data with a high co-efficient of variation (STORM et al., 2012). This occurs because there is plenty of variable that needs to be under control, and a huge variation among the animals. Additionally, there is no research that proves that the animal’s behavior is not changed with the use of equipment. Although, it was developed mainly for grazing cattle, this method requires high contact with the animals, and this could somehow disrupt them when they are not used to the presence of humans or their handling (HILL et
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al., 2016). Considering that the data from the SF6 tracer method could
also be used to develop the mathematical models and the predictions that will be compilated to the national inventory, the needed data is based upon realistic and reliable real-life statistics (IPCC, 2006). This means the maximum control of the aleatory variables could influence the results. In other words, in animal research, animals should behave normally, since the results are coming from other variables rather than animal behavior (POOLE, 1997). For this reason, a period of habituation with animals, is recommended before we research them. Thus, they can adapt to new conditions imposed upon them, avoiding jeopardizing their welfare and their productive performance (BROUCEK et al., 2017), such as the research results per se.
3.5 Influences that factor in dairy cow’s behavioral changes in research
When dairy cows are subjected to research, they are exposed to changes and conditions that are unusual from their normal routine. For instance, they are transferred from one housing system to another (ECKELKAMP et al., 2014; ENRIQUEZ-HIDALGO et al., 2017); they receive irregular handling (JOHNS; PATT; HILLMANN, 2015); their social environment changes (CACIOPPO et al., 2011; COSTA; VON KEYSERLINGK; WEARY, 2016), they have distinct different hygienic conditions in their housing (CHEN et al., 2017), they try different diets (DEVRIES; VON KEYSERLINGK; WEARY, 2004), and they are needed to test equipment and techniques for accepted validation (GOLDHAWK, BEAUCHEMIN, 2013; AMBRIZ-VILCHIS et al., 2015).
Dairy cows are very adaptable animals (JAGO, KERRISK, 2011). However, in certain situations and conditions imposed upon them, they, for a short period of time, require longer periods to adapt, thus causing behavioral changes. Behavioral changes are the natural response to environmental stimuli and provides the inside track to the physiological and psychological status of the cows (O’DRISCOLL et al., 2019). The cow’s main visible behavioral changes have been associated with lying behavior and ingestive behavior (eating and ruminating). Lying behavior, for instance, can be an indication of limited space (POPESCU et al., 2013), illness (SEPÜLVEDA-VARAS; WEARY; VON KEYSERLINGK, 2014), hunger and low food availability (O’DRISCOLL et al., 2015). Once lying is a priority for the cows (TUCKER et al., 2009) and is have been associated with the cow
