تعیین دما و pH بهینه رشد باکتری‌های تجزیه کننده مواد لیگنوسلولزی جدا شده از دستگاه گوارش موریانه‌ و اثر آن‌ها بر قابلیت هضم و فراسنجه‌های تخمیر آزمایشگاهی برخی پسماندهای کشاورزی

نوع مقاله : مقاله پژوهشی

نویسندگان

1 گروه علوم دامی دانشگاه لرستان

2 گروه علوم دامی دانشگاه کشاورزی و منابع طبیعی رامین خوزستان

3 گروه زیست‌شناسی دانشگاه شهید چمران اهواز

4 مؤسسه تحقیقات علوم دامی کشور

چکیده

زمینه مطالعاتی: روده موریانه‌ها حاوی انواع مختلفی از میکروارگانیسم‌های تجزیه کننده مواد لیگنوسلولزی است که امکان استفاده از آنها در بهبود ارزش تغذیه‌ای پسماندهای زراعی گامی بزرگ جهت پوشش دادن کمبود خوراک دام است. هدف:  این پژوهش به منظور تعیین دما و pH بهینه رشد باکتری‌های همزیست تجزیه کننده مواد لیگنوسلولزی جدا شده از دستگاه گوارش موریانه‌ها و بررسی قابلیت هضم و فراسنجه‌های تخمیر آزمایشگاهی کاه گندم و سرشاخه خرمای فرآوری شده با ایزوله‌های مذکور انجام شد. روش­کار: برای این منظور، از سه گونه باکتری Bacillus licheniformis ، Ochrobactrum intermedium و Microbacterium paludicola، استفاده گردید که باکتری‌های مذکور توسط کشت محتویات روده موریانه‌ها در محیط‌های کشت حاوی انواع مختلفی از لیگنین و مواد لیگنوسلولزی جداسازی شده بودند. منحنی‌های رشد ایزوله‌ها در محیط کشت نوترینت براث به دست آمد. و سپس کاه گندم و سرشاخه خرما توسط جدایه‌‌های مذکور عمل‌آوری شدند. نتایج: رشد بهینه باکتری B. licheniformisدر دمای 40 درجه سانتی‌گراد بود، اما در دو گونه  O. intermedium و M. paludicola رشد بهینه در دمای 37 درجه سانتی‌گراد مشاهده گردید. رشد بهینه هر سه سویه در pH برابر با 7 اتفاق افتاد. پس از عمل‌آوری، برای هر دو سوبسترای کاه گندم و سرشاخه خرما بیشترین حجم گاز تولیدی و قابلیت هضم ماده آلی در تیمار فرآوری شده توسط باکتری B. licheniformisو کمترین در تیمار شاهد مشاهده گردید (05/0P<). قابلیت هضم ماده خشک، سوبسترای تجزیه شده حقیقی، انرژی قابل متابولیسم، ضریب تفکیک، pH و غلظت نیتروژن آمونیاکی در هر دو سوبسترا تحت تأثیر تیمارهای آزمایشی قرار نگرفت. نتیجه‌گیری نهایی: نتایج این تحقیق نشان داد که عمل‌آوری کاه گندم و سرشاخه خرما توسط باکتری‌های جداسازی شده سبب بهبود فراسنجه‌های تخمیر شکمبه در شرایط برون‌تنی گردید.

کلیدواژه‌ها


عنوان مقاله [English]

Determination of optimum temperature and pH for lignocellulosic materials-degrading bacteria isolated from termite gut and their effect on the digestibility and in vitro fermentation parameters of some agricultural by-products

نویسندگان [English]

  • A Azizi 1
  • T Mohammadabadi 2
  • H Motamadi 3
  • M Chaji 2
  • H Fazaeli 4
Introduction: There is a shortage of animal feed and water resources in several developing countries. Thus, proper use of agro industrial by products is a useful means to overcome this problem. Agricultural by products can use as animal feeds, but because of high lignin and low protein content, their digestibility and palatability is low. Several processing method has been used to improving nutritive value of such materials (Rahal et al. 1997 and Fazaeli et al. 2004). Recently, biological delignification of lignocellulosic material has been considered as an alternative approach (Fazaeli et al. 2004). Termites gut contains different kinds of lignocellulose degrading microbes, which the possibility of their use in improving the nutritive value of agricultural by-products is a major step in compensation of animal feed shortage.
Material and methods: The present study was conducted to determine the optimum temperature and pH for growth of symbiont bacteria degrading lignocellulosic material isolated from termite gut, and to investigate their effects on the digestibility and in vitro fermentation parameters of wheat straw (WS) and date leaf (DL). For this purpose, three bacteria including Bacillus licheniformis, Ochrobactrum intermedium and Microbacterium paludicola were used,which isolated by culture of termite guts contents in mediums containing different types of lignin and lignocellulosic materials. Sterile basal media was used as a medium for isolation process and contained the following ingredients per litre: 7.0 g K2HPO4, 3.0 g KH2PO4, 1.0 g (NH4)2SO4, 0.1 g MgSO4·7H2O (Borji 2003). Growth curves of isolated bacteria for optimum temperature and pH were obtained using nutrient broth medium. Three temperatures of 30, 37 and 40 oC and four pH of 5, 6, 7 and 8 were evaluated. After that, WS and DL were processed with these isolates in media 9 (M9) medium with following composition (per litre): 6.2 g Na2HPO4, 3.0 g KH2PO4, 0.5 g NaCl, 1.0 g NH4Cl (Kato et al. 1998). After processing, residues were collected, dried and used for in vitro gas production experiment according to Makkar (2010). For this purpose, rumen liquor was collected from two ruminally fistulated cows. Animals were maintained on a 60% hay and 40% concentrate diet according to their requirements for two weeks. Rumen contents were collected from the ventral sac of rumen before morning feeding. A completely randomized design was used to determine the optimum temperature and pH for bacterial growth as well as effects of different bacteria on in vitro gas production and fermentation parameters of treated substrates. Data were analyzed by General Linear Model (GLM) procedures of SAS (SAS Institute, 2001),
Results and discussion:All of isolates were capable of grow at all of examined temperatures (i.e., 30, 37 and 40 oC). The optimum growth of B. licheniformis occurred at 40 oC, but the optimum growth of O. intermedium and M. paludicola was observed at 37 oC. All of isolates grew at different pH ranges (i.e., 5, 6, 7 and 8) and their optimum growth was observed at neutral pH of 7.0. At this neutral pH, highest growth for B. licheniformis, O. intermedium and M. paludicola was obtained after 30, 34 and 36 h of incubation, respectively. After processing, in both WS and DL, highest and lowest gas production and organic matter (OM) digestibility were observed by B. licheniformis and control treatment, respectively (P<0.05). In vitro dry matter (DM) digestibility, truly digested substrate, metabolizable energy, partitioning factor, pH and ammonia nitrogen concentration were not affected by the experimental treatments. Factors such as temperature, pH, atmosphere gas composition, bacterial type, composition of media culture, number of bacteria used and end products bacteria and their interaction has an important effect on growth rate of isolates (Mohan and Manuselis 1995). Similar to our results, by isolation of Bacillus sphaericus, Ochrobactrum anthropi and Enterobacter cloacae from termite gut, Borji (2003) reported that their optimum temperature and pH were observed at 25-37 oC and 7-8, respectively. Other researchers also reported similar results (Giroux et al. 1988 and Yang et al. 1995). Totally, obtaining optimizing pH and temperature of isolated bacteria for processing agricultural by-products can maximize their nutritive value. Improving OM digestibility and fermentation parameters in both substrates was maybe due to effect of isolates on changing lignin structure and loosening its bounds with carbohydrate compounds (VanSoet 2004). Consistent to results of present study, borji (2003) indicated that inoculation of wheat straw and barley straw with different bacteria isolated from termite gut increased their DM digestibility compared to control treatment. Increasing DM and OM digestibility and volume of gas production in WS than DL was probably because of lower lignin and higher carbohydrate content in the former (Azizi-Shotorkhoft et al. 2016).     
Conclusion:Results of present study showed that optimum growth for Ochrobactrum intermedium and Microbacterium paludicola isolated from termite gut was observed at 37 oC while it was 40 oC for Bacillus licheniformis. Optimum pH for growth of all of isolates was 7. Processing wheat straw and date leaf with bacteria, especially B. licheniformis improved rumen fermentation parameters in vitro. Research for upgrading the nutritive value of by products with ligninolytic bacteria isolated from termite gut is in infancy. Thus, further works in this field of study is needed.