Bacterial species and evolution: Theoretical and practical perspectives

A discussion of the species problem in modern evolutionary biology serves as the point of departure for an exploration of how the basic science aspects of this problem relate to efforts to map bacterial diversity for practical pursuits—for prospecting among the bacteria for useful genes and gene-products. Out of a confusing array of species concepts, the Cohesion Species Concept seems the most appropriate and useful for analyzing bacterial diversity. Techniques of allozyme analysis and DNA fingerprinting can be used to put this concept into practice to map bacterial genetic diversity, though the concept requires minor modification to encompass cases of complete asexuality. Examples from studies of phenetically definedBacillus species provide very partial maps of genetic population structure. A major conclusion is that such maps frequently reveal deep genetic subdivision within the phenetically defined specles; divisions that in some cases are clearly distinct genetic species. Knowledge of such subdivisions is bound to make prospecting within bacterial diversity more effective. Under the general concept of genetic cohesion a hypothetical framework for thinking about the full range of species conditions that might exist among bacteria is developed and the consequences of each such model for species delineation, and species identification are discussed. Modes of bacterial evolution, and a theory of bacterial speciation with and without genetic recombination, are examined. The essay concludes with thoughts about prospects for very extensive mapping of bacterial diversity in the service of future efforts to find useful products. In this context, evolutionary biology becomes the handmaiden of important industrial activities. A few examples of past success in commercializing bacterial gene-products from species ofBacillus and a few other bacteria are reviewed.     

Thermal ecotypes of amphi-Atlantic algae. I. Algae of Arctic to cold-temperate distribution (Chaetomorpha melagonium,Devaleraea ramentacea andPhycodrys rubens)

Three species of Arctic to cold-temperate amphi-Atlantic algae, all occurring also in the North Pacific, were tested for growth and/or survival at temperatures of −20 to 30°C. When isolates from both western and eastern Atlantic shores were tested side-by-side, it was found that thermal ecotypes may occur in such Arctic algae.Chaetomorpha melagonium was the most eurythermal of the 3 species. Isolates of this alga were alike in temperature tolerance and growth rate but Icelandic plants were more sensitive to the lethal temperature of 25°C than were more southerly isolates from both east and west. With regard toDevaleraea ramentacea, one Canadian isolate grew extraordinarily well at −2 and 0°C, and all tolerated temperatures 2–3°C higher than the lethal limit (18–20°C) of isolates from Europe. ConcerningPhycodrys rubens, both eastern and western isolates died at 20°C but European plants tolerated the lethal high temperature longer, were more sensitive to freezing, and attained more rapid growth at optimal temperatures. The intertidal species,C. melagonium andD. ramentacea, both survived freezing at −5 and −20°C, at least for short time periods.C. melagonium was more susceptible thanD. ramentacea to desiccation. Patterns of thermal tolerance may provide insight into the evolutionary history of seaweed species.     

Characterization of recombinant C1 inhibitor P1 variants.

Twelve human C1 inhibitor P1 variants were constructed by site-directed mutagenesis of the codon for arginine 444 and were expressed in COS-1 cells to analyze the functional properties. The ability to bind to target proteases, as well as potential substrate-like behavior, was investigated with radioimmunoassays. The P1-Lys variant retained binding capacity toward C1s, plasmin, and kallikrein. In addition, complex formation with C1s was detected for P1-Asn and P1-His. All other P1 substitutions resulted in C1 inhibitor variants that neither complexed with nor were inactivated by C1s, kallikrein, beta-factor XIIa, or plasmin. Electrophoretic studies confirmed that P1-Lys and P1-His can form sodium dodecyl sulfate-resistant complexes with C1s. In contrast, the C1s-P1-Asn complex dissociated upon addition of sodium dodecyl sulfate. Kinetic experiments by the method of progress curves generated association rate constants (kon) with C1s of 4.2 x 10(4) M-1 s-1 for recombinant wild-type C1 inhibitor and 1.7 x 10(4) M-1 s-1 for P1-Lys. For P1-Asn and P1-His, kon was decreased approximately 100-fold. The results from inhibition experiments were compatible with a model of reversible inhibition, although the observed dissociation rate for wild-type C1 inhibitor is too low (1-2 x 10(-6) s-1) to be physiologically relevant. The overall inhibition constant (Ki) was estimated to be 0.03 nM. With P1-Asn, reversible inhibition could be demonstrated directly upon dilution of preformed complexes; the observed dissociation rate constant was 3.2 x 10(-4) s-1; and Ki increased to approximately 380 nM. These findings are discussed in relation to inhibitor specificity and inhibition mechanism.     

Sequencing and analysis of the cos region of the lactococcal bacteriophage c2

The cohesive termini of the DNA genome of the lactococcal bacteriophage c2 were directly sequenced and appeared to be complementary, non-symmetrical, 9-nucleotide single-stranded, 3 extended DNAs, with the following sequence: 5-GTTAGGCTT-3 3-CAATCCGAA-5. DNA located on either side of the cohesive ends was sequenced and several repeats and a region with the potential for a DNA bend were found. Previously sequenced cos regions of 13 other bacteriophages were also examined for similar sequence features. All of the bacteriophages from gram-positive hosts had 3 extended DNA termini, in contrast to the bacteriophages from gram-negative hosts, which had 5 extended DNA termini. All bacteriophages had a region of dyad symmetry close to the cohesive termini. A 7.3 kb DNA fragment of the c2 genome containing the cos sequences was cloned; transduction experiments demonstrated that these cloned sequences could act as a substrate for packaging enzymes of phage c2.     

K-ATP opener-mediated attenuation of spontaneous bladder contractions in ligature-intact,partial bladder outlet obstructed rats

Symptoms of urinary frequency and urgency secondary to benign prostatic obstruction are common in elderly men. In many patients, these symptoms correspond to the urodynamic finding of involuntary detrusor contractions during filling cystometry (i.e., detrusor instability). Spontaneous non-voiding contractions during filling can be modeled in animals by subchronic, partial urethral obstruction. However, many investigators remove the obstructive ligature a few days prior to cystometrical evaluation (which may not be an ideal representation of the clinical situation where obstruction is still present), and all perform cystometry within 3 days post-bladder catheterization surgery (i.e., while considerable wound healing is present). In the current study, we evaluated the effects, after oral dosing, of three structurally diverse ATP-sensitive potassium channel openers (KCOs) on spontaneous contractions secondary to obstruction in rats with an intact obstructive ligature at the time of testing and 2 weeks post-bladder catheterization. ZD6169, WAY-133537 and a novel dihydropyridine KCO, A-278637, all significantly decreased spontaneous bladder contractions at 30 min post-dosing (p.o.). However, only ZD6169 (10 micromol/kg) and A-278637 (3 micromol/kg) attenuated such bladder contractions at doses that did not concurrently, significantly affect mean arterial blood pressure and heart rate. These data confirm the efficacy of KCOs to inhibit unstable contractions in obstructed rats, and they further demonstrate the positive effect of a novel, bladder-selective KCO, A-278637, in an animal model with potentially less artifact than in previous such models.     

Residue incorporation depth is a controlling factor of earthworm‐induced nitrous oxide emissions

Earthworms can increase nitrous oxide (N₂O) emissions, particularly in no‐tillage systems where earthworms are abundant. Here, we study the effect of residue incorporation depth on earthworm‐induced N₂O emissions. We hypothesized that cumulative N₂O emissions decrease with residue incorporation depth, because (i) increased water filled pore space (WFPS) in deeper soil layers leads to higher denitrification rates as well as more complete denitrification; and (ii) the longer upward diffusion path increases N₂O reduction to N₂. Two 84‐day laboratory mesocosm experiments were conducted. First, we manually incorporated maize (Zea mays L.) residue at different soil depths (incorporation experiment). Second, ¹³C‐enriched maize residue was applied to the soil surface and anecic species Lumbricus terrestris (L.) and epigeic species Lumbricus rubellus (Hoffmeister) were confined to different soil depths (earthworm experiment). Residue incorporation depth affected cumulative N₂O emissions in both experiments (P < 0.001). In the incorporation experiment, N₂O emissions decreased from 4.91 mg N₂O–N kg^?1 soil (surface application) to 2.71 mg N₂O–N kg^?1 soil (40–50 cm incorporation). In the earthworm experiment, N₂O emissions from L. terrestris decreased from 3.87 mg N₂O–N kg^?1 soil (confined to 0–10 cm) to 2.01 mg N₂O–N kg^?1 soil (confined to 0–30 cm). Both experimental setups resulted in dissimilar WFPS profiles that affected N₂O dynamics. We also found significant differences in residue C recovery in soil organic matter between L. terrestris (28–41%) and L. rubellus (56%). We conclude that (i) N₂O emissions decrease with residue incorporation depth, although this effect was complicated by dissimilar WFPS profiles; and (ii) larger residue C incorporation by L. rubellus than L. terrestris indicates that earthworm species differ in their C stabilization potential. Our findings underline the importance of studying earthworm diversity in the context of greenhouse gas emissions from agro‐ecosystems.     

Suppression and recovery of estrous behavior in Syrian hamsters after changes in metabolic fuel availability

A reduction in the availability of oxidizable metabolic fuels inhibits reproduction. Forty-eight hours of metabolic fuel deprivation inhibits estrous behavior in ovariectomized, steroid-treated Syrian hamsters, but little is known about the time course of this inhibition. Likewise, refeeding reverses deprivation-induced suppression, but the rate of recovery has not been examined. In two experiments we determined 1) the rate at which estrous behavior declines in hamsters treated with metabolic inhibitors and 2) how rapidly sexual receptivity is restored when hamsters are refed after a 48-h fast. We also measured circulating levels of leptin and insulin in an attempt to determine their relationship to the inhibition and restoration of estrous behavior. More than 24 h of metabolic inhibitor administration were required to inhibit lordosis, whereas only 6 h of refeeding were sufficient to restore the display of sexual receptivity to normal levels. Neither plasma insulin nor leptin levels paralleled the changes in estrous behavior. We concluded that 1) suppression of estrous behavior occurs more slowly than recovery after a fast and 2) changes in circulating leptin and insulin probably do not have a critical role in these behavioral changes.