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Aflatoxins are naturally occurring toxic chemical substances that are produced by fungal species called Aspergillus flavus. The toxic substances are secondary metabolites, which contaminate groundnut while growing in the field and also post-harvest. Drought stress is one of the factors that contribute to increased aflatoxin levels in groundnut during field production. This study was conducted in a screen house at ICRISAT-Chitedze Agricultural Research Station, Malawi to investigate the effects of drought on aflatoxin contamination and A.flavus population in the soil. Four drought stress levels; prolonged (4 weeks), min (3 weeks), mild (2 weeks) and no drought were imposed on five groundnut varieties at pod filling stage. Soil samples were collected from each plot four times; at planting, beginning of drought, end of drought and at harvest. Aflatoxin levels in groundnut grain samples were estimated by use of neogenstrips read with mobile assay tablet reader. Population densities of A.flavus in soil samples collected from the plots were estimated using serial dilutions plated on the selective media, modified dichloran Rose Bengal (MDRB) and quantify A.flavus within 3 days after incubation at 37°C.The results showed that there were significant differences in aflatoxin contamination between drought stress levels (p = 0.011). High aflatoxin contamination was observed under prolonged drought (22.0 ppb) compared to and no drought treatment (1.5 ppb). None of the varieties used in the study showed either resistance or susceptibility to aflatoxin contamination under drought or adequate soil moisture. The results also showed that there were significant differences in A.flavus population at drought period and harvesting time and the mean population of A.flavus in prolonged drought at end of stress and harvesting were 8511 and 6044 cfu/g of soil respectively. It was concluded that drought contribute to aflatoxin contamination in groundnut, and also increased the A.flavus population in soil and also at harvesting.

Peanut (Arachis hypogaea L.) growers must balance complex interactions among cultivars, planting dates, environmental and physiological stresses during the growing season, and weather conditions at harvest when determining when to dig peanut. Ten field experiments were conducted in North Carolina from 1994 through 1996 to determine the influence of digging date on pod yield and gross return of virginia-type peanut. Beginning in mid- to late September, the cultivars NC 9, NC 10C, NCV-11, VA-C 92R, AgraTech (AT) VC-1, and NC 12C were dug on four dates approximately 7 d apart. Considerable variation in pod yield and gross return was noted among cultivars and experiments. Delaying digging increased pod yield and gross return in some but not all experiments. Greater variation in pod yield and gross return was observed for NC 10C than for AT VC-1 when compared across digging dates. Pod yield and gross return for NC9, NC V-11, VA-C 92R, and NC 12C were intermediate between NC 10C and AT VC-1. Of the cultivars evaluated, yield and gross return of AT VC-1 were the most stable over digging dates. These data suggest that growers should evaluate maturity of peanut in individual fields for each cultivar when determining when to dig. These data also suggest that factors other than maturity impact pod yield and gross return.

Experiments were conducted to compare barnyardgrass control and rice injury and yield with emulsifiable concentrate and dry flowable formulations of propanil as single or repeat applications with crop oil concentrate, methylated seed oil, a blend of organosilicone surfactant and methylated seed oil or conventional nonionic surfactant, and organosilicone surfactant. Two applications of propanil were more effective in controlling barnyardgrass than a single application. The emulsifiable concentrate formulation of propanil controlled barnyardgrass more effectively than the dry flowable formulation in some but not all experiments. Differences in barnyardgrass control with propanil as influenced by adjuvants were minor and inconsistent. The most consistent barnyardgrass control and the highest rice yields were obtained with repeat applications of the emulsifiable concentrate formulation of propanil.

Light harvesting and utilization by chloroplasts located near the adaxial vs the abaxial surface of sun and shade leaves were examined by fluorometry in two herbaceous perennials that differed in their anatomy and leaf inclination. Leaves of Thermopsis montana had well-developed palisade and spongy mesophyll whereas the photosynthetic tissue of Smilacina stellata consisted of spongy mesophyll only. Leaf orientation depended upon the irradiance during leaf development. When grown under low-light levels, leaves of S. stellata and T. montana were nearly horizontal, whereas under high-light levels, S. stellata leaves and T. montana leaves were inclined 60° and 30°, respectively. Leaf inclination increased the amount of light that was intercepted by the lower leaf surfaces and affected the photosynthetic properties of the chloroplasts located near the abaxial leaf surface. The slowest rates of quinone pool reduction and reoxidation were found in chloroplasts located near the adaxial leaf surface of T. montana plants grown under high light, indicating large quinone pools in these chloroplasts. Chloroplasts near the abaxial surface of low-light leaves had lower light utilization capacities as shown by photochemical quenching measurements. The amount of photosystem II (PSII) down regulation, measured from each leaf surface, was also found to be influenced by irradiance and leaf inclination. The greatest difference between down regulation monitored from the adaxial vs abaxial surfaces was found in plants with horizontal leaves. Different energy dissipation mechanisms may be employed by the two species. Values for down regulation in S. stellata were 2-3 times higher than those in T. montana, while the portion of the PSII population which was found to be Q B nonreducing was 4-6 times lower in high light S. stellata leaves than in T. montana. All values of Stern-Volmer type nonphotochemical quenching (NPQ) from S. stellata leaves were similar when quenching analysis was performed at actinic irradiances that were higher than the irradiance to which the leaf surface was exposed during growth. In contrast, with T. montana, NPQ values from the abaxial leaf surface were up to 45% higher than those from the adaxial leaf surface regardless of growth conditions. The observed differences in chloroplast properties between species and between the adaxial and abaxial leaf surfaces may depend upon a complex interaction among light, leaf anatomy and leaf inclination.

The subalpine environment of the Medicine Bow Mountains, Wyoming (USA) is characterized by frost events throughout the summer. As a result, growing season length is difficult to define in terms of frost-free days. For example, such a definition would have limited the 1993 growing season for many plants to only 5 days. The frequency of summertime frost events appeared particularly dependent on net losses of infrared energy to a cold night sky (radiation frost). These radiative losses to the night sky were strongly dependent on microsite sky exposure and leaf characteristics as well as meteorological conditions. During the summer of 1993, air temperatures at a height of 8 cm at the center of a subalpine meadow were below 0°C on 26 of 67 nights, while leaves of a broadleaf species ( Erigeron peregrinus ) frosted on 41 nights compared to 25 nights for a coniferous sapling ( Abies lasiocarpa ). In E. peregrinus , a strong association occurred between the frequency and duration of freezing episodes and the amount of sky exposure, with plants sheltered by forest canopy experiencing frost only 10% to 40% as often as those in the center of the meadow. At exposed locations, longwave radiation minima from the night sky were strongly correlated with the occurrence of leaf temperature minima. Thus, microsite sky exposure as well as the variation in longwave radiation from the atmosphere may have important implications for estimating growing season length and plant distribution patterns at higher elevations. Growth habit also influenced the occurrence of frost because plant height spanned a substantial vertical gradient in air temperature above the soil surface. This was particularly true on nights with low sky infrared radiation and low wind. On calm, clear nights, the coldest leaf temperatures ( T leaf ≈ −11°C with T air ≈ −6°C) occurred because of the combined effects of low sky infrared radiation and low air temperature at plant heights. However, even on relatively windy nights with air temperature above freezing, net radiative losses to clear skies resulted in leaf frost. Flower temperatures of both E. peregrinus and Polygonum bistortoides tended to be significantly warmer (≈ 2°C) than leaf temperatures due to their greater height above the ground. Thus, leaf size, plant height, microsite exposure, and daily variation in sky infrared radiation all have important influences on the determination of frost frequency, duration, and intensity, as well as the length of the growing season.

Terminology characterizing differences in ear growth habit and prolificacy is commonly used by seed companies to relate corn (Zea mays L.) hybrid yield potential to varying plant populations. Little information is available to document the significance of various ear response types associated with commercially available Corn Belt hybrids. The objective of this study was to assess hybrid x plant population interactions of commercial corn hybrids differing in ear growth habit and prolificacy. Field experiments were conducted in 11 environments in Ohio during 1990 and 1991. Four hybrids (LH119 x LH51, B73 x LH38, LH132 x DM2, and LH123 x LH93) chosen to represent different responses to plant population associated with ear type were compared at three plant populations (16 000, 24 000, and 32 000 plants/acre). Grain yields, ears per plant, and stalk lodging varied with hybrid and plant population in both years. The semiprolific hybrid and the fixed and flex single ear hybrids exhibited similar yield response to plant population across a wide range of environmental conditions. In 1990, grain yields were greatest for the flex (LH51 x LH119) and fixed (B73 x LH38) single ear hybrids and semiprolific (LH123 x LH93) hybrid at 32 000 plants/acre and the prolific (LH132 x DM2) hybrid at 24 000 plants/acre; whereas in 1991, under drier, hotter conditions, yields for all four hybrids were greatest at 24 000 plants/acre. Prolificacy was most pronounced in LH132 x DM2 and at the low plant population; it was least evident in LH119 x LH51 and at the high plant population. Stalk lodging was greatest in the prolific and semiprolific hybrids and at the high plant population; it was negligible in the fixed single ear hybrid and at the low plant population. Although the prolific hybrid exhibited relatively high yields under stress in 1991, its greater predisposition to lodging limits its potential use in Eastern Corn Belt environments where diseases and weather conditions make stalk quality a major factor in hybrid selection. The relatively minor role that hybrid x population interactions played in determining grain yield compared with environment, hybrid, and plant population main effects suggests that hybrid differences in ear growth habit and prolificacy are of limited importance in determining optimum plant populations.

Objective: Late proptosis is an unusual complication of orbital floor fracture repair. We report 12 cases and propose a classification scheme and differential diagnosis for this rare complication. Design: Retrospective multi-institutional case review. Setting: Two tertiary referral centers. Results: We report 12 cases of late proptosis following orbital floor fracture repair. Causes of late proptosis include peri-implant inflammation, capsular hemorrhage, gelatin film cyst, implant infection, sino-orbital fistula, intraorbital sinus mucocele, and carotidcavernous fistula. We describe the treatment of each case individually; in most cases, the implant was removed. All 12 patients had a satisfactory outcome after treatment. Conclusions: Proptosis presenting 2 months or more after orbital floor fracture is rare. We classify the origin of late proptosis into three groups: implant, sinus, and neurovascular. We describe the diagnosis and management of this unusual complication. (Arch Otolaryngol Head Neck Surg. 1995;121:649-652)

Neanderthal pelvic morphology is not well understood, despite the recent find and analysis of the Kebara 2 pelvis. Many of the proposed hypotheses focus on the possible need for a larger birth canal. A previously unexplored aspect involves possible direct obstetric implications of bone robusticity and density. These characteristics ocan affect obstetrics in modern humans, especially the molding of the neonate's head during parturition: engineering studies have shown that denser neonate cranial bones undergo less deformation, and thicker (more robust) cranial bones would also be expected to deform less during the birth process. These bone characteristics may also result in a less flexible birth canal. Thus, more robust or denser bones could result in the need for a larger birth canal or a smaller neonate head, due to decreased flexibility. Examples from modern populations are discussed and the conclusions applied to Neanderthals, who are known to have had high bone robusticity and may have had high bone density, given their heavy musculature. (A positive association between muscle mass and bone density has been observed repeatedly in modern humans.) We conclude that bone robusticity and density may have obstetrical implications for Neanderthals, with particular importance for neonate head molding during birth.

The light interception capabilities of individual conifer needles are governed by their cross-sectional geometry and their orientation to sunlight. Leaf cross sections typical of conifer tree species were modeled to quantify the interception of direct sunlight over a range of incident light angles. The needle shapes exhibited by Abies nordmanniana Spach, Picea asperata Master, Pinus cembra L., P. monophylla TorrFrém., and P. sylvestris L. were selected because they are representative of the range of geometric shapes found in conifer tree species. Calculated light interception values were compared to corresponding predictions for a laminar broadleaf. Estimates of carbon gain were derived from computed incident light integrated over the leaf cross section and a representative curve of conifer photosynthetic response to light. Flat leaf cross sections (e.g., Abies nordmanniana) with high surface area to volume ratios (6) intercepted more light per unit area at high angles of incidence than thick leaves. Thick leaves (e.g., Pinus cembra) intercepted more light at low angles of incidence than at high angles of incidence. Needles of Pinus monophylla had no angular dependence for light interception because of their circular cross section. Large differences in estimated CO(2) assimilation occurred among the species, especially when CO(2) uptake was expressed on a unit volume basis. A maximum uptake of 67.9 mmol CO(2) m(-3) s(-1) was predicted for A. nordmanniana compared to a minimum of 39.7 mmol m(-3) s(-1) for P. monophylla. A greater angular dependence occurred for estimates of CO(2) uptake than for estimates of light interception.

The barbel was originally a fish with a relatively restricted range in the British Isles. However, despite the general environmental degradation of many British rivers, and in contrast with some native freshwater fishes, the barbel is now more widely distributed than it was a century ago. It may even be more abundant numerically.

An apparently new species of Clostridium was isolated from mud samples obtained from a stagnant lake near Athens, Georgia. These organisms were able to utilize inorganic pyrophosphate as a source of energy for growth, a phenomenon representing the simplest ATP-generating system in the biological world.

Heating of Streptococcus faecium NRC 1261 at 55 C for 15 min produced injury. The injured population was sensitive to 2.5% NaCl but regained its tolerance when incubated in a recovery medium. Addition of penicillin and actinomycin D to the recovery medium showed that during recovery no cell division occurred and that this recovery was linked to RNA synthesis. Analysis of the fatty acid composition of uninjured and recovered cells by gas liquid chromatography showed an increased concentration of saturated fatty acids in recovered cells. A concurrent increase in resistance to heat shock at 55 C and salt tolerance in these recovered cells occurred with the change in fatty acid composition. It is suggested that a change in membrane structure as reflected by the modified fatty acid profile is related to an increased thermal resistance and salt tolerance.

The acetylene-reduction assay was used for in situ and laboratory assessment of biological nitrogen fixation in the acidic, waterlogged, muskeg ecosystem of the southern James Bay area, in the region of Moosonee, Ontario. In situ assays and subsequent laboratory experimentation revealed that nitrogenase activity was predominately a function of the activities of heterocystic blue-green bacteria associated with surface water, with the phyllosphere of mosses, and with at least one lichen, a species of Peltigera. No such in situ activity was detected in the subsurface organic material, even when such material was amended with glucose. However, under laboratory conditions at 20 °C, nitrogenase activity was evident in the subsurface layers after an extended lag and was shown to be higher under anaerobic than under aerobic conditions, to have an optimum temperature range extending about a mean of 20 °C, and to be stimulated by glucose. This potential for subsurface nitrogen fixation proved to be related to the presence of microorganisms existing in anaerobic microsites within the organic layers and no microorganisms capable of fixation could be detected under aerobic incubation.

The present study evaluates possible effects of adrenal catecholamines, released by splanchnic nerve stimulation, on adrenal medullary blood flow (MQ) and adrenal catecholamine secretion (CS). Twelve pentobarbital-anesthetized mongrel dogs were subjected to three identical splanchnic nerve stimulations (5 V, 20 Hz, for 3 min) at 30-min intervals, and MQ (radiolabeled microsphere technique) and CS (high-performance liquid chromatography) were measured before and during each nerve stimulation. Animals were assigned to one of three groups and administered either saline, pindolol (1 and 4 mg/kg), or prazosin (1 and 4 mg/kg) before the second and third nerve stimulation, respectively. In the saline control group, each nerve stimulation resulted in similar increases in MQ and CS. Pindolol attenuated nerve stimulation-induced increases in MQ and CS by 50%, but had no effect on medullary catecholamine concentration. Prazosin augmented nerve stimulation-induced MQ, CS, and medullary catecholamine concentration by 35%. These data suggest that adrenal adrenergic receptors modulate elicited CS and mediate changes in adrenal medullary vascular tone.

The transport of succinate was studied in bacteroids of an effective, streptomycin-resistant strain (GF160) of Rhizobium leguminosarum. High levels of succinate transport occurred, and the kinetics, specificity, and sensitivity to metabolic inhibitors were similar to those previously described for free-living cells. The symbiotic properties of two transposon (Tn5)-mediated C4-dicarboxylate transport mutants (strains GF31 and GF252) were determined. Strain GF31 formed ineffective nodules, and bacteroids from these nodules showed no succinate transport activity. Strain GF252 formed partially effective nodules, and bacteroids from these nodules showed about 50% of the succinate transport activity of the parent bacteroids. Another dicarboxylic acid transport mutant (Dct-), strain GFS5, isolated after N-methyl-N'-nitro-N-nitrosoguanidine mutagenesis, formed ineffective nodules. The ability to form ineffective nodules in strains GF31 and GFS5 was shown to correlate with the Dct- phenotype. The data indicate that the presence of a functional C4-dicarboxylic acid transport system is essential for N2 fixation to occur in pea nodules.

Large-scale movements of fish within a complex freshwater system such as the Norfolk Broads can give rise to erroneous sampling data if the extent of these movements is not fully appreciated. The Norfolk Broads habitat is briefly described, and data are presented showing that mean estimates of standing crop in the broads vary from

Recent data indicate that the slow-growing, non-acid-producing root nodule bacteria of leguminous plants should be separated from the fast-growing, acid-producing strains and placed in a new genus. The separation is warranted by numerical taxonomy, deoxyribonucleic acid base ratio determinations, nucleic acid hybridization, ribosomal ribonucleic acid cistron similarities, serology, composition of extracellular gum, carbohydrate utilization and metabolism, bacteriophage and antibiotic susceptibilities, protein composition, and types of intracellular inclusion bodies in the bacteroid forms. The name proposed for the new genus is Bradyrhizobium. The type species of the genus is B. japonicum (Buchanan 1980) comb. nov. (basonym: Rhizobium japonicum Buchanan 1980), the type strain of which is ATCC 10324.

Adult roach, bream and their presumed F1 hybrid from an Anglian Water reservoir were identified on the basis of morphological and meristic characteristics. The hybrid was clearly intermediate. Four hybrid breeding crosses were induced to spawn by hypophysis. A bream × roach cross (female named first) failed to produce fertile eggs, whereas F1 hybrid × roach, roach × F1 hybrid and F1 hybrid × F1 hybrid all produced fry. Fertility (defined as survival of eggs to hatching) was high for the F1 hybrid × roach back-cross (56%) but low for the others (

Hemorrhagic hypotension produces an increase in adrenal medullary blood flow and a decrease in adrenal cortical blood flow. To determine whether changes in adrenal blood flow during hemorrhage are neurally mediated, we compared blood flow responses following adrenal denervation (splanchnic nerve section) with changes in the contralateral, neurally intact adrenal. Blood pressure was reduced and maintained at 60 mmHg for 25 min by hemorrhage into a pressurized bottle system. Adrenal cortical blood flow decreased to 50% of control with hemorrhage in both the intact and denervated adrenal. Adrenal medullary blood flow increased to four times control levels at 15 and 25 min posthemorrhage in the intact adrenal, but was reduced to 50% of control at 3, 5, and 10 min posthemorrhage in the denervated adrenal. In a separate group of dogs, the greater splanchnic nerve on one side was electrically stimulated at 2, 5, or 15 Hz (n = 4 each group) for 40 min. Adrenal medullary blood flow increased 5- to 10-fold in the stimulated adrenal but was unchanged in the contralateral, nonstimulated adrenal. Adrenal cortical blood flow was not affected by nerve stimulation. We conclude that activity of the splanchnic nerve profoundly affects adrenal medullary vessels but not adrenal cortical vessels and mediates the observed increase in adrenal medullary blood flow during hemorrhagic hypotension.

SUMMARYThe proteins and the amino acid profile of the rumen fungi Neocallimastix frontalis and Piromonas communis have been examined and the in vitro digestibility of the vegetative growth determined.The mean true protein content of N. frontalis was 24% of the dry weight and 30% for P. communis, which has a lower chitin content. 50% of the protein extracted from cells and examined by HPLC was in the 200000 MW range and 40% in the 50000 MW range. Polyacrylamide gel electrophoresis after treatment with sodium dodecylsulphate and 2-mercaptoethanol indicated that the native proteins were composed of smaller units in the MW range 25000–50000.The amino acid profiles of the two fungi were similar and compared favourably with those of casein and fraction 1 protein of lucerne.

Two recent reports described the isolation of derivatives of a Rhizobium trifolii strain that had gained the ability to nodulate Glycine max and Vigna radiata and that had demonstrated altered patterns of carbon source utilization, free-living nitrogen fixation, and hydrogen uptake. More extensive characterization of these strains now supports the conclusion that these strains are R. japonicum and are not derived from the putative parent R. trifolii .

Arranged in descending order of nitrogen-fixing (acetylene-reducing) potential the sites examined were mesic meadow and peat polygon troughs (equal rank), transition zone between mesic meadow and gravel ridge, gravel ridge, polar dessert, and peat polygon tops. The dominant nitrogen-fixing microorganisms, as in other Arctic areas, were blue-green bacteria, especially those epiphytic on Arctic mosses. The epiphytic association exhibited an optimum temperature for fixation of 20 degrees C. Other bacteria potentially able to fix nitrogen were present in the soils examined but their activity was severely restricted by low soil temperatures and lack of readily utilizable energy sources. These bacteria included members of the genera Klebsiella (the most numerous), Bacillus, Clostridium, and Beijerinckia (scarce). Also present at many of the sites was an unidentified yellow-pigmented fixer which was not Mycobacterium flavum. All fixers were psychotrophic rather than psychrophilic, having an optimum temperature greater than 20 degrees C but capable of slow growth at 5 degrees C or lower. The rate of acetylene reduction by the epiphytic system increased with the number of successive exposures to acetylene, a phenomenon of some significance in any calculations designed to measure the amount of nitrogen fixed in certain ecosystems.

Medicago laciniata, an annual leguminous plant of Saharo-Sindian origin, is particularly refractory to root nodulation by most strains ofRhizobium meliloti. Using a series of such bacterial strains belonging to the 8 groups of Brockwell and Hely, and a variety of environmental conditions, it was noted that several normally non-nodulating strains (at 20°C) produced ineffective nodules at root temperatures of 24°C to 28°C. Nodulation at 20°C failed to occur in the presence of a wide variety of test compounds and physical conditions. No phytoalexins or anti-Rhizobium growth inhibitors were isolated from inoculated root tissue at any temperature. Temperature shift experiments indicated no infection of the root hairs at 20°C, and infection threads produced at the permissive root temperature failed to elongate after transfer to 20°C. However, if meristematic activity had been initiated in the inner root-cortical cells as a result of infection thread penetration at 28°C, no blockage of nodule maturation occurred upon subsequent transfer to 20°C root temperature. Nodules produced at 28°C were completely devoid of nitrogenase activity, although the apical (but not the distal) regions contained normal-appearing bacteriods, surrounded by enclosing membranes, and possessed a fully functional leghaemoglobin. A shortage of metabolic energy did not appear to be involved in the ineffective response. A hypothesis to explain the nodulation phenomenon observed was based on the observation in the roots of 2 factors present at 20°C but not at 28°C.

Ammonium chloride was not utilized by three strains of Rhizobium meliloti as the sole source of nitrogen in a sucrose medium, unless either amino or certain non-nitrogenous carboxylic acids were also present. This was also essentially true for the utilization of nitrate, nitrite, purines, and pyrimidines, all of which are potentially able to form ammonia. These results may be interpreted on the assumption that washed cells of alfalfa – sweet clover rhizobia require, for growth initiation in a nitrogen-free medium, either preformed amino acids or compounds such as ammonia and certain carboxylic acids from which amino acids can be synthesized. Since α-ketoglutarate was extremely active in promoting growth in a medium containing ammonium chloride it was implied that the ammonia may be fixed by L-glutamic acid dehydrogenase activity, especially since this particular enzyme was located in these organisms. No aspartase activity could be demonstrated. The ineffective strain differed from the effective strains in that it was unable to use purines or pyrimidines as accessory nitrogen sources in amino acid media. This was a result of strain variation and it was not coupled with the state of ineffectiveness itself. A synthetic medium has been formulated for further growth studies on washed Rhizobium cells and for investigations on auxotrophic mutants of these bacteria.

Jordan , D. C. (Ontario Agricultural College, Guelph, Canada), I. Grinyer, and W. H. Coulter . Electron microscopy of infection threads and bacteria in young root nodules of Medicago sativa . J. Bacteriol. 86: 125–137. 1963.—Ultrathin sections of alfalfa nodules ( Medicago sativa L.) revealed that initially the infection thread consisted of a naked mucoid strand, containing imbedded bacteria, migrating between the walls of the cortical cells of the host plant. At certain locations, the plant cell walls were forced into a funnel-shaped structure which elongated to form a tubular sheath containing bacterial cells and gum. Lateral and terminal vesicles formed on the ensheathed infection thread, and these burst to liberate the bacteria into the host cell cytoplasm, the process being similar to that proposed by certain of the earlier workers. In some cases, the ensheathed thread migrated entirely across a host cell without rupturing, and, in this case, the tip of the thread fused into the host cell wall opposite the point of entry. The bacteria, after their release from the thread, became bacteroidal, and ultimately each bacterial cell became surrounded by a double-layered membrane. It is suggested that this enclosing membrane may be derived from the tubules of the endoplasmic reticulum, although the possibility of the formation of these membranes de novo as a protective mechanism of the host plant cannot entirely be disregarded.

A simple and rapid procedure is described for the isolation of plasma membrane from the gram-negative bacterium Rhizobium meliloti. The method involves the mechanical disruption of cells followed by differential and density gradient centrifugation. Cell wall, intermediate, and cell membrane fractions were isolated. Electron microscopy indicated no attachment of the membrane to the wall although some membrane material was trapped within the wall fragments. The intermediate fraction consisted of both wall and membrane material and the membrane fraction appeared to be free of wall material. Biochemical data strongly supported the electron microscopic evidence. Chemical analyses of the uncontaminated membranes indicated that they possessed a composition consistent with other bacterial membranes.

Four distinct types of colonial mutation were obtained by X-ray irradiation of a parasitic strain of alfalfa Rhizobium. Two types were procured from an effective strain. Colonial morphology remained stable throughout serial transfer on artificial medium, but changed considerably after one plant passage. Three variants derived from the parasitic parent were found to be efficient in nitrogen fixation when tested by sterile plant growth procedures and the Virtanen technique. This efficiency increased during the course of two plant passages. Colonial variation and effectivity changes occurred independently of one another.

No intracellular infection threads were observed in ultrathin sections of young root nodules of lupine, although nodule bacteria could be found in the intercellular spaces between the root cortical cells. Evidence suggests that in certain instances the plant cell walls can be disrupted locally, allowing the bacteria to pass into cytoplasm of the host cell. The spread of infection may be initiated in this manner and extended by division of infected cells. No plant-produced enclosing membranes were present around bacteria in the intercellular spaces but such structures developed after the bacteria had entered the plant cell. Although the origin of these membranes is debatable, in the present work it appeared that they were formed de novo, perhaps in a manner akin to the development of the cell plate during cell division. Most of the bacterial cells possessed a wide subwall space lying between the bacterial cell wall and plasma membrane. Discontinuities present in the latter membrane may account for the ribosome-like material found in the subwall space.

The biochemical mechanism of action of viomycin on R. meliloti involved an inhibition of protein synthesis. Adenine and amino acid incorporation into total nucleic acid and cell wall respectively was relatively unaffected. The antibiotic increased the level of 14C in the intracellular pool and "nucleic acids" fraction of cells fed 14C-amino acids but caused no cellular lysis. Mono- and di-valent cations protected cells from growth inhibition by viomycin.A viomycin-resistant mutant was similar to its sensitive parent in rate of uptake of glucose and glutamic acid by both growing and resting cells and in the rate of leakage when preloaded cells were suspended in buffered saline. Viomycin resistance did not appear to be episome-mediated or involve an extracellular degradative enzyme. The ability of TES buffer to eliminate viomycin resistance, coupled with the sensitivity to viomycin of cell-free protein-synthesizing systems derived from both sensitive and resistant cells indicates that resistance is probably the result of a selective reduction in antibiotic permeability.

The Virtanen technique was used to detect effective and ineffective rhizobia. The rhizobia studied varied widely in their action towards different varieties of the same host species. Maximum leghemoglobin concentrations occurred just prior to blossoming in field legumes, but much earlier in greenhouse legumes cultivated under conditions of this experiment. The nodules of the ineffective strains of legume bacteria differed from those produced by effective strains in color, size, distribution, rate of destruction of the bacteroid area, and presence or absence of rodlike bacterial forms. Experimental evidence is presented to show that a supposedly parasitic strain of alfalfa organism can exist.

Cells of Rhizobium meliloti contained an intracellular pool of amino acids. The passage of C14-histidine or C14-glutamate into this pool apparently occurred against a concentration gradient and was energy dependent, being inhibited by azide and dinitrophenol. An internal bound form of C14, containing a number of radioactive amino acids, was synthesized at the expense of radioactive pool components. This bound C14consisted predominantly of several proteins whose syntheses were inhibited by chloramphenicol. Added histidine was not incorporated directly into this protein since the protein histidine was nonradioactive. When C14-labelled cells were suspended in glucose-containing basal medium there was an efflux of pool amino acids into the extracellular environment. This leakage presumably was the result of simple diffusion since it was energy independent, occurred with the concentration gradient, and was appreciable at 5 °C. Rapid leakage occurred at 30 °C in the presence and absence of dinitrophenol for an initial period of 6 hours, after which time growth occurred in the absence of this inhibitor. During growth leakage ceased, and a considerable quantity of material passed back into the cells and was incorporated into a bound form. In the presence of dinitrophenol no growth occurred, and the outflow continued after the 6th hour, but at a much reduced rate. X irradiation increased the efflux of pool constituents, but the reason for this is conjectural.

Under the experimental conditions used, amino acids played a very important part in the growth initiation of washed cells of alfalfa – sweet clover rhizobia. There were, however, distinct differences in utilization, both among genetically related mutants, and among other cultures when compared before and after plant passage. None of 15 vitamins, purines, and pyrimidines was able to initiate growth and hence these rhizobia are able to synthesize these compounds when a readily utilizable nitrogen source is present. Because of this fact, the stimulation of these bacteria by yeast extract is probably due, primarily, to the amino acid content. No strain was found able to concentrate free amino acids intracellularly or, when grown in lysine or tyrosine media, to excrete additional amino acids. No differences were found among effective, ineffective, or parasitic rhizobia in biochemical requirements.

Under clearly defined conditions one-step acquisition of viomycin resistance by a normally effective strain of Rhizobium meliloti resulted in one-step acquisition of ineffectiveness in nitrogen fixation, which probably occurred with a one-gene change in the R. meliloti genome. Two-step mutants retained their ability to produce root nodules but such nodules also were ineffective. Increased sensitivity of the viomycin-resistant mutants to glycine and D-alanine was not noted. Bacteroids were not seen in nodules formed by the viomycin-resistant mutants on their homologous host plant. Nitrogenase activity was not detected, by acetylene reduction, in detached ineffective nodules, whereas effective nodules formed 10.6 μmoles of ethylene per hour per gram of nodules. Growth of the effective parent strain in a low concentration of viomycin resulted in elongation and swelling of the cells so that they appeared as artificially produced bacteroids. Viomycin-resistant mutants did not undergo this transformation. Antigens could be readily extracted by hot- and cold-saline extraction of wet packed cells of both resistant and sensitive cultures but antigenic differences, which may have indicated cell wall differences, were not noted.

Growth of Lotus nodule bacteria (Strain LC 296) was temporarily suppressed in a yeast-extract mineral-salts medium containing 2,4-DB at 50 and 100 µg/ml. However, the 2,4-DB concentration needed to completely suppress the growth of the bacteria appeared to be above 500 µg/ml. When added during the logarithmic phase of growth 2,4-DB at 50 and 100 µg/ml did not inhibit the growth of the cells, so that the growth-inhibiting effect of the herbicide apparently was directed primarily against lag phase cells. Pre-incubation of rhizobia in a medium containing 2,4-DB at concentrations up to 10 µg/ml did not affect the capacity of the bacteria to effectively nodulate their host plant. Attempts to induce the rhizobia to utilize 2,4-DB as a sole carbon source failed. Nevertheless, when theLotus nodule bacteria (LC 296) were grown in a medium containing 2,4-DB-1-C14, C14O2 was released. Similar results were obtained with other strains of the Lotus rhizobia and other species of Rhizobium. C14O2 evolution was also demonstrated when radioactive 2,4-DB was added to both intact (excised) and crushed trefoil nodules. The results indicate that the nodule bacteria are capable of degrading 2,4-DB; although the mechanism is still obscure it does not appear to be via cleavage of the ether link.

Field trials carried out in 1965 and 1966 showed that 2,4-DB, alone or in combination with dalapon, reduced nodulation and tended to decrease the efficiency of nitrogen fixation in birdsfoot trefoil. Dalapon appeared to enhance the inhibitory action of 2,4-DB on nodulation. No obvious cytological differences could be detected in the nodules or in the isolated bacteroids of field-treated and untreated plants. Under growth chamber conditions, 2,4-DB drastically reduced trefoil growth and nodulation particularly in treatments where the herbicide came directly in contact with the plants. It appears that the reduction in nodulation and nitrogen fixation is a result of plant damage and abnormal root growth caused by 2,4-DB application.

Studies to determine the effect of various concentrations of sodium nitrate and ammonium chloride on the ability of Rhizobium meliloti to infect and fix nitrogen in three varieties of alfalfa indicated that nodulation was inhibited at high levels of combined nitrogen. High nitrate concentrations inhibited nodulation to a greater extent than high ammonium concentrations. A small amount of combined nitrogen, however, appeared to promote nodulation. Inoculated plants grown in nitrogen levels which might be encountered in the field consistently contained more nitrogen than uninoculated plants under the same treatment. There was comparatively little difference in the nitrogen content or behaviour of the varieties of alfalfa tested.

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Identification of Rhizobium and Bradyrhizobium strains and especially of indigenous isolates continues to be one of the major difficulties associated with competition studies. Because there is no universally accepted method, the method of choice depends on preference, experience, and equipment. Here, an agar plate technique was used to distinguish strains and field isolates of Rhizobium leguminosarum biovar trifolii to provide a basis for identifying nodule occupants in further competition studies. A rapid plate technique, based on differential growth characteristics, complements other techniques such as serological reactions, particularly when antisera cross-react with nonhomologous strains. The technique involves culturing strains and isolates on chrome azurol S agar. Although similar responses were observed among some strains, the response was highly reproducible and was considered an ideal complementary technique used in conjunction with serological procedures. Strains with similar responses could often be differentiated by varying media components, such as the source of carbon.

Atmospheric oxygen is a potent inhibitor of photosynthesis in plants utilizing the C3 pathway. This inhibition is mediated by ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco), in that CO2 and O2 are mutually competitive substrates on the enzyme for RuBP. Oxygenation of RuBP yields glycolate-P, which is subsequently metabolized to CO2 and PGA in photorespiration. The interaction between photosynthesis and photorespiration is determined by the kinetic constants of Rubisco and the chloroplast CO2 and O2 concentrations as described in Eq. 1 (Laing et al. 1974)

In three ineffective associations between Medicago sativa (alfalfa) and Rhizobium meliloti the initial stages of nodule formation, resulting in the release of the bacteria into the host cells, were found to follow the normal pattern of nodule development. In nodule tissue formed by two laboratory-produced ineffective mutants, a rapid disintegration of the invading bacteria was observed to occur shortly after the release of the bacteria into the plant cells. The disintegrating bacteria were in intimate association with large amounts of rough endoplasmic reticulum (ER). An increase in the number of mitochondria occurred at this stage as well and the peripheries of the plant cells were often lined with starch granules. Only occasionally was the stage of enclosing-membrane formation reached. In the third ineffective association, a naturally occurring one, the bacteria were transformed into the nitrogen-fixing or bacteroidal forms and were surrounded by enclosing membranes. Dissolution of the bacteria occurred at a slightly later stage in this association and was again accompanied by a buildup of rough ER. Evidence is presented to suggest that the plant response, as characterized by this ER buildup in these ineffective associations, was a manifestation of nitrogen starvation.

Yellow blotch disease of the oyster mushroom ( Pleurotus ostreatus ) was first observed in a commercial mushroom farm in California in 1983. The disease, caused by Pseudomonas agarici , is characterized by primordia, with yellow droplets on their surface, which become stunted, yellow to orange, and deformed as they mature.

Several members in two families were found to have a pericentric inversion of chromosome 19. A review of four previous cases, together with those reported here, suggests that inversion of chromosome 19 is not related to the phenotypic features of the probands. Furthermore, there has been no report of an affected subject resulting from a duplication deficiency product of inverted chromosome 19 among the offspring of inversion heterozygotes. The suggested association of aneuploidy in the inversion carriers is also discussed.

Bacteroids of Rhizobium leguminosarum were formed rapidly in media containing 0.35% yeast extract, but not in media containing 0.1% yeast extract. Bacteroid production in the former media could be prevented by increasing the oxygen tension, suggesting that the limited oxygen supply within root nodules may be the principal factor initiating the natural formation of bacteroids. The artificially produced bacteroids were shorter and possessed a more irregular external morphology than bacteroids extracted from red root nodules of field pea, although the internal organization of both types of bacteroids was similar.During the development of bacteroids in laboratory media there was a rapid decrease in viable count and a concurrent increase in nephelos, although this increase was at a reduced rate compared to that exhibited by the normal small rod forms of rhizobia. These data indicate an inhibition of cellular division and a continued synthesis of at least certain protoplasmic constituents. In this connection the capacity to synthesize nucleic acids was similar in both artificially produced bacteroids and the small, rod types of the test organism, although some minor differences were found with bacteroids in situ within the nodules when nodule age varied.All types of bacteroids, whether they were within the nodule, had been extracted from the nodule, or were produced in laboratory media, suffered an impairment in the synthesis of protein and cell wall. Therefore, in several important respects, artificially produced and naturally produced bacteroids are physiologically similar. The alteration in wall-synthesizing capacity and the possible formation of limited amounts of abnormal wall material incapable of employing certain ions, such as Mg2+, for maintenance of rigidity is a likely cause of both the inhibition of cellular division and of the abnormal external morphology of these rather unusual cells.

Mutation to viomycin-resistance in Rhizobium meliloti R21 resulted in an accumulation of phosphatidylcholine and phosphatidylethanolamine in the cell wall. Resistance to viomycin decreased when the excess lipid was removed by EDTA or when its synthesis was prevented by growth of normally resistant cells at 40 °C. Microelectrophoretic data showed binding of viomycin to the cell surface and it is proposed that the mechanism of resistance to viomycin is an immobilization of the antibiotic in the surface layers of the cell as a result of combination with phospholipid.

0736 01 1. A mutant strain of Salmonella thompson (S. thompson/r), with an increased resistance to gamma radiation, did not appear to possess a DNA repair system, active in a “non-growth” medium, similar to that reported for Micrococcus radiodurans and Escherichia coli B/r. 2. DNA strand breaks occurred to a similar degree in both the sensitive parent strain and the resistant mutant strain during exposure to gamma radiation. 3. The DNA could be reconstituted in both strains, but only when the cells were incubated in a growth-promoting medium after irradiation. 4. Results indicated that inactivation of S. thompson by gamma radiation was not the result of the production of DNA single-strand breaks per se, and the resistance of the mutant strain, therefore, did not appear to be directly related to the ability to repair such breaks.