Sodium, an Obligate Growth Requirement for Predominant Rumen Bacteria

Sodium is an obligate growth requirement for most currently recognized predominant species of rumen bacteria. The isoosmotic deletion of Na(+) from a nutritionally adequate defined medium completely eliminated growth of most species. Growth yields and rates were both a function of Na(+) concentration for Na(+)-requiring species, and Na(+) could not be replaced by Rb(+), Li(+), or Cs(+) when these ions were substituted for Na(+) at a concentration equivalent to an Na(+) concentration that supported abundant growth. Li(+), Cs(+), or Rb(+) was toxic at an Na(+)-replacing concentration (15 mM) but not at a K(+)-replacing concentration (0.65 mM). K(+) was also an obligate growth requirement for rumen bacteria in media containing Na(+) and K(+) as major monovalent cations, but K(+) could be replaced, for most species, by Rb(+). The quantities of Na(+) that support rapid and abundant growth of Na(+)-requiring rumen bacteria show that these organisms are slight halophiles. A growth requirement for Na(+) appears more frequent among nonmarine bacteria than has been previously believed.     

Carbon Dioxide as an Essential Requirement for Cultured Sycamore Cells

Carbon dioxide (optimum concentration c. 1.0%) is essential to the initiation of the growth in suspension culture or on agar plates of cultured sycamore cells. By effective flushing of the cultures with CO₂-free air it is possible to demonstrate this requirement with initial cell densities up to 50 × 10³ cells ml^?1. This growth-promoting activity of carbon dioxide is not related to any effect it may have on the pH of the culture medium. The cells fix applied carbon dioxide into organic and amino acids but attempts to replace the carbon dioxide requirement by non-toxic levels of organic or amino acids have not been successful.     

Isotope Fractionation in Photosynthetic Bacteria during Carbon Dioxide Assimilation

The δ _PDB¹³C values have been determined for the cellular constituents and metabolic intermediates of autotrophically grown Chromatium vinosum. The isotopic composition of the HCO₃^- in the medium and the carbon isotopic composition of the bacterial cells change with the growth of the culture. The δ _PDB¹³C value of the HCO₃^- in the media changes from an initial value of −6.6‰ to +8.1‰ after 10 days of bacterial growth and the δ _PDB¹³C value of the bacterial cells change from −37.5‰ to −29.2‰ in the same period. The amount of carbon isotope fractionation during the synthesis of hexoses by the photoassimilation of CO₂ has a range of −15.5‰ at time zero to −22.0‰ after 10 days. This range of fractionation compares to the range of carbon isotope fractionation for the synthesis of sugars from CO₂ by ribulose 1,5-diphosphate carboxylase and the Calvin cycle.     

Effect of Carbon Dioxide on Growth of Meat Spoilage Bacteria

The ability of CO₂ to inhibit respiration and growth of representative strains of seven species of meat spoilage bacteria was examined. Enterobacter and Microbacterium thermosphactum were unaffected by CO₂. Both respiration and growth of the other species were inhibited. With four of the species (fluorescent and nonfluorescent Pseudomonas, Alteromonas putrefaciens, and Yersinia enterocolitica), the inhibition pattern in a complex medium was similar, and inhibition was incomplete and reached a maximum level at comparatively low concentrations of CO₂. With Acinetobacter, inhibition continued to increase with increasing CO₂ concentration. The degree of inhibition with a constant concentration of CO₂ in solution increased with decreasing temperature for all CO₂-susceptible species except the nonfluorescent Pseudomonas. Anaerobic growth of CO₂-susceptible facultative anaerobes was unaffected by CO₂.     

Lysis of Viable Rumen Bacteria in Bovine Rumen Fluid

Streptococcus bovis and Butyrivibrio sp. were labeled with thymidine-methyl-(3)H, washed, and resuspended in rumen fluid or rumen fluid fractions obtained from Holstein and Jersey cows fed alfalfa hay once daily. Factors affecting the lytic activity found in untreated rumen fluid were examined. Day to day variation and differences before and after feeding were observed for the same cow. There were also differences between cows on the same day. For a given rumen fluid, the rate of release of label was roughly proportional to the number of labeled cells present over a 100-fold range in concentration. Removal of protozoa largely abolished the lytic action of fresh rumen fluid for S. bovis, but some soluble lytic activity remained. Mixed rumen protozoa added to media containing labeled S. bovis caused label to appear in solution. In a sample of rumen fluid containing 4.3 x 10(4) protozoa/ml 5.2% of the S. bovis population were destroyed by protozoa per hr. The mean rate of destruction for 12 runs on whole rumen fluid was 8.7% per hr with a standard deviation of 6.05. Parallel experiments with Butyrivibrio indicated that soluble lytic factors were more important for this organism. They could be destroyed by autoclaving and were generated when viable rumen bacteria were resuspended in autoclaved rumen fluid. The lysis of S. bovis and Butyrivibrio, at equal cell densities, by mixed rumen protozoa was compared in 30% rumen fluid media, and Butyrivibrio appeared to be more readily lysed than S. bovis.     

Requirement for Carbon Dioxide for Nonsymbiotic Expression of Rhizobium japonicum Nitrogenase Activity

The expression and maintenance of nitrogenase (C₂H₂) activity in growing, microaerobic liquid cultures of Rhizobium japonicum 3I1b110 was found to be stringently dependent on the sustained supply of CO₂. This requirement for CO₂ appeared to exceed the basal requirement for growth and was not related to effects on pH.     

Apparent Reassimilation of Respiratory Carbon Dioxide by Different Plant Species

Differences among species in respiration rates in CO₂-free air, in light and dark, were studied using the standard leaf chamber technique and the infrared carbon dioxide analyzer. Photosynthesis, transpiration and respiration were measured. In all species studied, rates of respiration were considerably higher in dark than in light. This effect was assumed to be due to reassimilation of the respiratory CO₂. A resistance analogy model was derived to account for the apparent differences in internal recycling of CO₂ among species; the differences were correlated with differences in maximum photosynthetic rates in normal air and optimal conditions (P₃₁₀) and with internal resistances to CO₂ diffusion (r_k). Species with high P₃₁₀ and low r_k appear to reassimilate all the endogenous CO₂, whereas other species with lower P₃₁₀ and higher r_k appear to reassimilate only a part of their respiratory CO₂. Experiments with the photosynthetic inhibitor, 3-(3,4-dichlorophcnyl)-l,l-dimethyl urea (DCMU), indicated that species with zero respiration in CO₂-free air and light release respiratory CO₂ when photosynthesis is inhibited. It is concluded that the CO₂ released in the presence of DCMU represents respiratory CO₂ which recycles to photosynthesis under normal conditions.     

Presence of Rhapidosomes in Various Species of Bacteria and Their Morphological Characteristics

Rod-shaped structures have been observed in cells of Pseudomonas, Photobacterium, Proteus, and Saprospira by use of the negative-contrast stain. These structures, referred to as rhapidosomes, appear to be normal components of these cells. Other bacteria including Escherichia, Salmonella, Shigella, Klebsiella, Micrococcus, Bacillus, Mycobacterium, Rhodospirillum, and Hydrogenomonas genera failed to reveal these structures. The rhapidosomes of Saprospira were found to consist of two rods, one encasing a narrower, longer structure. In contrast, the rhapidosomes of Pseudomonas, Proteus, and Photobacterium were without the rigid inner structure, but were occasionally seen filled with a homogeneous material as observed by the negative stain. Ultrathin sections of Pseudomonas cells indicate that these rhapidosomes are embedded within or are in close association with the nucleoplasm.     

瘤胃纤维素降解细菌的研究进展

反刍动物瘤胃是自然界中最有效的纤维素降解系统，其纤维素降解能力主要源于寄居于其中的纤维素降解细菌、真菌和原虫。其中，瘤胃纤维素降解细菌因数量庞大、种类繁多以及代谢途径丰富，在木质纤维素降解及利用方面发挥着重要作用。本文综述了国内外瘤胃纤维素降解细菌的种类，分析了瘤胃纤维素降解细菌的特性；阐述了瘤胃纤维素降解细菌通过纤维小体对纤维素的降解过程，以及瘤胃微生物之间的相互作用和相互制约关系；简述宏组学技术在开发新纤维素降解菌和新纤维素酶方面的应用，旨在为进一步研究纤维素降解细菌的降解机理，开发新的纤维素菌种和酶资源提供新的思路。     

Cell Envelope Morphology of Rumen Bacteria

The cell walls of three species of rumen bacteria (Bacteroides ruminicola, Bacteroides succinogenes, and Megasphaera elsdenii) were studied by a variety of morphological methods. Although all the cells studied were gram-negative and had typical cytoplasmic membranes and outer membranes, great variation was observed in the thickness of their peptidoglycan layers. Megasphaera elsdenii evidenced a phenomenally thick peptidoglycan layer whose participation in septum formation was very clearly seen. All species studied have cell wall "coats" external to the outer membrane. The coat of Bacteroides ruminicola is composed of large (approximately 20 nm) globules that resemble the protein coats of other organisms, whereas the coat of Bacteroides succinogenes is a thin and irregular carbohydrate coat structure. Megasphaera elsdenii displays a very thick fibrillar carbohydrate coat that varies in thickness with the age of the cells. Because of the universality of extracellular coats among rumen bacteria we conclude that the production of these structures is a protective adaptation to life in this particular, highly competitive, environment.     

Comparison of Rumen Microbial Inhibition Resulting from Various Essential Oils Isolated from Relatively Unpalatable Plant Species

Essential oils were isolated from eight plant species which were relatively unpalatable to sheep and deer. The inhibitory potency of these essential oils upon sheep and deer rumen microorganisms was compared, in terms of total gas and volatile fatty acid (VFA) production, by use of an anaerobic manometric technique. Inhibitory effects of oils from the eight plant species may be placed in four groups: (i) essential oils from vinegar weed (Trichostema lanceoletum) and California bay (Umbellularia californica) inhibited rumen microbial activity most; (ii) lesser inhibition was exhibited by rosemary (Rosmarinus officinalis) and California mugwort (Artemisia douglasiana) oils, followed by (iii) blue-gum eucalyptus (Eucalyptus globulus) and sagebrush (Artemisia tridentata) oils; and (iv) oils from Douglas fir (Psuedotsuga menziesii) and Jerusalem oak (chenopodium botrys) resulted in the least inhibition, when 0.3 ml of each oil was used. A highly significant correlation coefficient (r = 0.98(**)) between total gas and VFA production indicated the validity of either method to measure the activity of rumen microorganisms. Our results are discussed in relation to the hypothesis that the selectivity and voluntary consumption of ruminants are related to the characteristic odor and antibacterial action of essential oils isolated from relatively unpalatable plant species.     

Hemicellulose-degrading Enzymes Synthesized by Rumen Bacteria

Over 100 bacterial cultures isolated from ovine rumen contents by enrichment techniques in polysaccharide-containing media were examined for the ability to degrade plant cell wall structural polysaccharides. Approximately two-thirds of the isolates retained were Gram negative and amongst the coccoid isolates diplococci predominated. Many of the rods examined were piliated and/or flagellated. Thirty of the more active xylan- and arabinan-degrading isolates were examined for both the production and activity of the principal polysaccharidase and glycosidase enzymes associated with plant cell wall polysaccharide hydrolysis, following culture in a hemicellulose containing growth medium. The wide range of glycosidase activities detected in these hemicellulolytic isolates was evidence for their rôle in the breakdown of dietary polysaccharides in the rumen.     

Effects of Carbon Dioxide on Coccidian Oocysts from 6 Host Species*

SYNOPSIS Activation of sporozoites in oocysts of Eimeria acervulina (chicken), E. intricata (sheep), and E. scabra (swine) occurred after pretreatment in aqueous 0.02 M cysteine hydrochloride under an atmosphere of CO₂, followed by incubation in a trypsin-bile mixture. Sporozoites of E. stiedae (rabbit), E. bilamellata (squirrel), and Isospora canis (dog) became activated when incubated in trypsin and bile with or without prior CO₂-pretreatment of oocysts; however, when CO₂-pretreatment was used, activation of these species in trypsin and bile was greatly enhanced. For E. acervulina, 12% of the oocysts were activated after 4 hr CO₂-pretreatment and 10 hr incubation in trypsin and bile at 43 C; higher temperatures or longer pretreatment times did not cause greater activation. Eimeria intricata oocysts became activated after 1 hr pretreatment and 10 hr incubation in trypsin and bile at 37, 39 or 41 C, respectively. The highest activation (31%) occurred after 20 hr pretreatment and 10 hr incubation in trypsin and bile at 41 C. Ninety percent of E. scabra oocysts contained active sporozoites after 1 hr CO₂-pretreatment and 10 hr incubation in trypsin and bile at 37 C. At 39 or 41 C, 100% activation occurred with this species after similar pretreatment and treatment periods. With E. bilamellata, 64% activation occurred in nonpretreated oocysts incubated 10 hr in trypsin and bile at 41 C, whereas 100% activation occurred if oocysts were pretreated with CO₂ for 1 hr before treatment with trypsin and bile. Thirty-one, 35, and 36% of CO₂-pretreated E. stiedae oocysts were activated after 1 hr incubation in trypsin and bile at 37, 39 or 41 C, respectively, whereas 1, 2, and 20% activation occurred in nonpretreated oocysts incubated at the same temperatures. Sporozoites in 99-100% of I. canis oocysts were activated after 10 hr treatment in trypsin and bile with or without 1 hr CO₂-pretreatment at 23, 37, 39 or 41 C.     

Medium Without Rumen Fluid for Nonselective Enumeration and Isolation of Rumen Bacteria

Colony counts which approximated those in a habitat-simulating, rumen fluid-agar medium (RFM) were obtained in medium 10, a medium identical to the RFM except for the replacement of rumen fluid with 1.5 x 10(-6)m hemin, 0.2% Trypticase, 0.05% yeast extract, and a 6.6 x 10(-2)m volatile fatty acid mixture qualitatively and quantitatively similar to that in rumen fluid. Single deletion of Trypticase, yeast extract, or the volatile fatty acid mixture from medium 10 significantly reduced colony counts. Colony counts were also reduced when medium 10 was modified to contain higher concentrations of Trypticase or volatile fatty acids. Significant differences were found between colony counts obtained from diluted rumen contents of animals fed a cracked corn-urea diet, and the colony counts obtained from animals fed either a cracked corn-soyean oil meal or an alfalfa hay-grain diet. Qualitative differences were found between the predominant bacterial strains isolated from rumen contents of animals fed cracked corn diets and strains isolated from animals fed alfalfa hay-grain. Regardless of differences in the predominant flora associated with diet, medium 10 and the RFM supported growth of similar bacterial populations. The results show that medium 10 is suitable for enumeration and isolation of many predominant rumen bacteria.     

Catabolism of Lysine by Mixed Rumen Bacteria

Metabolites arising from the catabolism of lysine by the mixed rumen bacteria were chromatographically examined by using radioactive lysine. After 6 hr incubation, 241 nmole/ ml of lysine was decomposed to give ether-soluble substances and CO₂ by the bacteria and 90 nmole/ml of lysine was incorporated unchanged into the bacteria. δ-Aminovalerate, cadaverine or pipecolate did not seem to be produced from lysine even after incubation of the bacteria with addition of those three amino compounds to trap besides lysine and radioactive lysine. Most of the ether-soluble substances produced from radioactive lysine was volatile fatty acids (VFAs). Fractionation of VFAs revealed that the peaks of butyric and acetic acids coincided with the strong radioactive peaks. Small amounts of radioactivities were detected in propionic acid peak and a peak assumed to be caproic acid. The rumen bacteria appeared to decompose much larger amounts of lysine than the rumen ciliate protozoa did.     

Utilization Modes of Inorganic Carbon for Photosynthesis in Various Species of Chlorella

Rates of CO₂ and HCC₃^– fixation in cells of various Chlorellaspecies in suspension were compared from the amounts of ¹⁴Cfixed during the 5 s after the injection of a solution containingonly ¹⁴CO₂ or H¹⁴CO₃^–. Results indicated that irrespectiveof the CO₂ concentration during growth, Chlorella vulgaris 11h and C. miniata mainly utilized CO₂, whereas C. vulgaris C-3,C. sp. K. and C. ellipsoidea took up HCO₃^– in additionto CO₂. Cells of C. pyrenoidosa that had been grown with 1.5%CO₂ (high-CO₂ cells) mainly utilized CO₂, whereas those grownwith air (low-CO₂ cells) utilized HCO₃^– in addition toCO₂. Cells that utilized HCO₃^– had carbonic anhydrase(CA) on their surfaces. The effects of Diamox and CA on the rates of CO₂ and HCO₃^–fixation are in accord with the inference that HCO₃^– wasutilized after conversion to CO₂ via the CA located on the cellsurface. CA was found in both the soluble and insoluble fractions;the CA on the cell surface was insoluble. Independent of the modes of utilization, the apparent K_m (NaHCO₃)for photosynthesis was much lower in low-CO₂ cells than in high-CO₂ones. The fact that the CA in the soluble fraction in C. vulgarisC-3 was closely correlated with the K_m(NaHCO₃) indicates thatsoluble CA lowers the K_m. ¹ Dedicated to the late Professor Joji Ashida, one of the foundersand first president of the Japanese Society of Plant Physiologists. ⁴ On leave from Research and Production Laboratory of Algology,Bulgarian Academy of Sciences, Sofia. (Received September 14, 1982; Accepted March 1, 1983)     

Inactivation of the Phosphoenolpyruvate-Dependent Phosphotransferase System in Various Species of Bacteria by Vinylglycolic Acid

The alkenoic hydroxyacid 2-hydroxy-3-butenoic acid (vinylglycolate) specifically inhibited the phosphotransferase system in a variety of bacteria while not affecting respiration-coupled transport systems.     

Functional Body Capacities under the Conditions of Various Concentrations of Carbon Dioxide and Oxygen

We determined a permissible ratio between carbon dioxide and oxygen concentrations during accidental situations. The experiments (n = 138, 10 h each) on the effect of various concentrations of carbon dioxide and oxygen in the inhaled air were conducted on male volunteers aged 20–40 years subjected to a special medical examination. All experiments were divided into five series: hypercapnia + normoxia, hypercapnia + hyperoxia, hypercapnia + hypoxia, normocapnia + hypoxia, and ambient air (control). The results showed that functional capacities of the body are less impaired under the conditions of hypercapnia combined with hyperoxia. Thus, in accidental situations associated with rapid accumulation of carbon dioxide in the atmosphere of airtight chambers, a synchronous increase in pO₂ to 220–230 torr can provide for the highest work capacity.     

Carbon Dioxide Assimilation and Methane Oxidation in Various Zones of the Rainbow Hydrothermal Field

Rates of carbon dioxide assimilation and methane oxidation were determined in various zones of the Rainbow Hydrothermal Field (36°N) of the Mid-Atlantic Ridge. In the plume above the hydrothermal field, anomalously high methane content was recorded, the microbial population density (up to 10⁵ cells/ml) was an order of magnitude higher than the background values, and the CO₂ assimilation rate varied from 0.01 to 1.1 g C/(l day). Based on the data on CO₂ assimilation, the production of organic carbon due to bacterial chemosynthesis in the plume was calculated to be 930 kg/day or 340 tons/year (about 29% of the organic carbon production in the photic zone). In the black smoke above active smokers, the microbial population density was as high as 10⁶ cells/ml, the rate of CO₂ assimilation made up 5–10 g C/(l day), the methane oxidation rate varied from 0.15 to 12.7 l/(l day), and the methane concentration ranged from 1.05 to 70.6 l/l. In bottom sediments enriched with sulfides, the rate of CO₂ assimilation was at least an order of magnitude higher than in oxidized metal-bearing sediments. At the base of an active construction, whitish sediment was found, which was characterized by a high methane content (92 l/dm³) and a high rate of methane oxidation (1.7 l/(dm³ day)).     

Pectin-fermenting Bacteria Isolated from the Bovine Rumen

Thirty-two strains of pectin-fermenting rumen bacteria were isolated from bovine rumen contents in a rumen fluid medium which contained pectin as the only added energy source. Based on differences in morphology and the Gram stain, 10 of these strains were selected for characterization. Two strains were identified as Lachnospira multiparus, four strains were identified as Butyrivbrio fibrisolvens, and three strains were identified as Bacteroides ruminicola. Characteristics of the remaining strain did not correspond with any previously described species. It was a gram-positive anaerobic coccus, 1.0 to 1.2 mum in diameter, and occurred primarily as single cells or diplococci. The strain fermented pectin rapidly but showed little or no growth on any other energy sources tested. The only detectable end products were acetic acid and gas, a portion of which was identified as hydrogen. Although the physiological characteristics of this organism differ markedly from other described species, it has been placed in the genus Peptostreptococcus on the basis of morphology, Gram stain, relations to oxygen, and the occurrence of cell division in only one plane. End products of fermentation are somewhat similar to those of the cellulolytic ruminococci. Eight previously characterized strains of cellulolytic bacteria isolated in nonselective media were unable to ferment pectin, whereas ten strains of hemicellulolytic rumen bacteria, eight of which were isolated with a xylan medium, showed considerable variation in this characteristic.