The influence of space and time on the evolution of altruistic defence: the case of ant slave rebellion.

How can antiparasite defence traits evolve even if they do not directly benefit their carriers? An example of such an indirect defence is rebellion of enslaved Temnothorax longispinosus ant workers against their social parasite Temnothorax americanus, a slavemaking ant. Ant slaves have been observed to kill their oppressors' offspring, a behaviour from which the sterile slaves cannot profit directly. Parasite brood killing could, however, reduce raiding pressure on related host colonies nearby. We analyse with extensive computer simulations for the Temnothorax slavemaker system under what conditions a hypothetical rebel allele could invade a host population, and in particular, how host-parasite dynamics and population structure influence the rebel allele's success. Exploring a wide range of model parameters, we only found a small number of parameter combinations for which kin selection or multilevel selection could allow a slave rebellion allele to spread in the host population. Furthermore, we did not detect any cases in which the reduction of raiding pressure in the close vicinity of the slavemaker nest would substantially contribute to the inclusive fitness of rebels. This suggests that slave rebellion is not costly and perhaps a side-effect of some other beneficial trait. In some of our simulations, however, even a costly rebellion allele could spread in the population. This was possible when host-parasite interactions led to a metapopulation dynamic with frequent local extinctions and recolonizations of demes by the offspring of few immigrants.

Oh sister, where art thou? Spatial population structure and the evolution of an altruistic defence trait.

The evolution of parasite virulence and host defences is affected by population structure. This effect has been confirmed in studies focusing on large spatial scales, whereas the importance of local structure is not well understood. Slavemaking ants are social parasites that exploit workers of another species to rear their offspring. Enslaved workers of the host species Temnothorax longispinosus have been found to exhibit an effective post-enslavement defence behaviour: enslaved workers were observed killing a large proportion of the parasites' offspring. As enslaved workers do not reproduce, they gain no direct fitness benefit from this 'rebellion' behaviour. However, there may be an indirect benefit: neighbouring host nests that are related to 'rebel' nests can benefit from a reduced raiding pressure, as a result of the reduction in parasite nest size due to the enslaved workers' killing behaviour. We use a simple mathematical model to examine whether the small-scale population structure of the host species could explain the evolution of this potentially altruistic defence trait against slavemaking ants. We find that this is the case if enslaved host workers are related to nearby host nests. In a population genetic study, we confirm that enslaved workers are, indeed, more closely related to host nests within the raiding range of their resident slavemaker nest, than to host nests outside the raiding range. This small-scale population structure seems to be a result of polydomy (e.g. the occupation of several nests in close proximity by a single colony) and could have enabled the evolution of 'rebellion' by kin selection.

Macro- and microgeographic genetic structure in an ant species with alternative reproductive tactics in sexuals.

The genetic structure of social insect populations is influenced by their social organization and dispersal modes. The ant Hypoponera opacior shows diverse reproductive behaviours with regular cycles of outbreeding via winged sexuals and inbreeding via within-nest mating wingless sexuals that reproduce by budding. This unusual life cycle should be reflected in the genetic population structure, and we studied this on different scales using microsatellites. On a macrogeographic scale, populations were considerably structured and migration rates within the Chiricahuas were higher than those in between mountain ranges. On a local scale, our analyses revealed population viscosity through dependent colony foundation and a high genetic diversity with a multicolonial structure. The latter was also evident from recognition trials revealing consistent aggression between non-nestmates. Within-nest matings led to high inbreeding coefficients. Finally, the observed seasonal changes in relatedness can be explained by variation in queen number and differential dispersal of the two reproductive morphs.

Acid ceramidase 1 expression correlates with a better prognosis in ER-positive breast cancer.

OBJECTIVES: Ceramide and sphingosine mediate response to cancer therapy, inhibit cell growth and induce apoptosis in vitro. Only a few clinical data about the impact of ceramide and sphingosine iny vivo are available. We investigated the relevance of ceramide- and sphingosine-generating enzymes in breast cancer (acid ceramidase 1 (ASAH1), ceramide synthases 4 (LASS4) and 6 (LASS6)) by means of gene expression analysis. METHODS: We analyzed differences in ASAH1, LASS4 and LASS6 on mRNA level between breast cancer subgroups using microarray data from 1581 tumor samples. RESULTS: High ASAH1, LASS4 and LASS6 expression correlates with pathohistological grading (p < 0.001) and estrogen receptor (ER) status (p < 0.001). High ASAH1 expression was associated with a larger tumor size >2 cm (p = 0.003), while high LASS6 expression was correlated with ErbB2 negativity (p < 0.001). In survival analysis, we detected a significant better prognosis of patients with higher ASAH1 expression (p = 0.002) in the ER-positive subgroup. In contrast, expression of LASS4 or LASS6 did not show any prognostic impact. In the multivariate analysis, only ASAH1 expression (p = 0.002), tumor size (p < 0.0001) and ErbB2 positivity (p = 0.041) remained significant. CONCLUSION: ASAH1 is an estrogen-dependent member of the sphingolipid metabolism, which might provide further prognostic information in ER-positive breast cancers.

The involvement of phosphorylation of myosin phosphatase targeting subunit 1 (MYPT1) and MYPT1 isoform expression in NO/cGMP mediated differential vasoregulation of cerebral arteries compared to systemic arteries.

AIM: Constitutive release of NO blunts intrinsic and stimulated contractile activity in cerebral arteries (CA). Here, we explored whether phosphorylation and expression levels of the PKG-sensitive, leucine zipper positive (LZ+ ) splice variants of the regulatory subunit of myosin phosphatase (MYPT1) are involved and whether its expression is associated with higher cGMP sensitivity. METHODS: Vascular contractility was investigated by wire myography. Phosphorylation of MYPT1 was determined by Western blotting. RESULTS: Constitutive phosphorylation of MYPT1-T696 and T853 was lower and that of S695 and S668 was higher in cerebral arteries from the circulus arteriosus (CA-w) than in femoral arteries (FA), while total MYPT1 expression was not different. In CA-w but not in FA, L-NAME lowered phosphorylation of S695/S668 and increased phosphorylation of T696/T853 and of MLC20 -S19, plus basal tone. The increase in basal tone was attenuated in CA-w and basilar arteries (BA) from heterozygous MYPT1-T696A/+ mice. Compared to FA, expression of the LZ+ -isoform was ~2-fold higher in CA-w coincident with a higher sensitivity to DEA-NONOate, cinaciguat and Y27632 in BA and 8-Br-cGMP (1 µmol/L) in pre-constricted (pCa 6.1) α-toxin permeabilized CAs. In contrast, 6-Bnz-cAMP (10 µmol/L) relaxed BA and FA similarly by ~80%. CONCLUSION: Our results indicate that (i) regulation of the intrinsic contractile activity in CA involves phosphorylation of MYPT1 at T696 and S695/S668, (ii) the higher NO/cGMP/PKG sensitivity of CAs can be ascribed to the higher expression level of the LZ+ -MYPT1 isoform and (iii) relaxation by cAMP/PKA pathway is less dependent on the expression level of the LZ+ splice variants of MYPT1.

Band-shape analysis and resolution of electronic spectra of pyridoxal phosphate and other 3-hydroxypyridine-4-aldehydes.

The electronic absorption spectra of individual ionic forms of pyridoxal phosphate and of a series of related aldehydes have been evaluated together with pKa values. Spectral resolution with lognormal curves has permitted the quantitative description of equilibria for hydration and tautomerization. Precise values of peak positions for both aldehyde and hydrate forms have been obtained. Measurements of temperature-induced changes in the spectra have provided additional information. Knowing the hydration ratios and stepwide acid dissociation constants, it is possible to evaluate microscopic acid dissociation constants for both the aldehyde and hydrate forms of the compounds.

Role of an active site residue analyzed by combination of mutagenesis and coenzyme analog.

Asp222 of aspartate aminotransferase is an active-site residue which interacts with the pyridine nitrogen of the coenzyme, pyridoxal 5'-phosphate (PLP). The roles of Asp222 in the catalytic mechanism of Escherichia coli aspartate aminotransferase have previously been explored by site-directed mutagenesis. These studies confirmed that a negatively charged residue at position 222 is essential for catalysis, but the reason for this remained speculative. In the present studies, the roles of Asp222 were clarified experimentally by analyzing the mutant D222A enzyme (Asp222 replaced by Ala) reconstituted with the coenzyme analog N(1)-methylated PLP (N-MePLP). Spectroscopic and kinetic analyses showed that Asp222 stabilizes the protonated N(1) of PLP, raising the pKa value of N(1) by more than five units, in the active site of AspAT. The positive charge at N(1) accelerates abstraction of the alpha-proton from the amino acid substrate, stabilizing the transition state by 1.4 to 4.5 kcal.mol-1 in the reaction with aspartate. X-ray crystallographic (2.0 A resolution) and CD spectroscopic studies suggest that the coenzyme analog is not held in a proper orientation within the active site of D222A (N-MePLP). This may account for the finding that the catalytic activity was recovered only partially by the reconstitution of D222A with N-MePLP. These results fully support the following postulated role of Asp222: the negative charge of Asp222 stabilizes the positive charge at N(1) of PLP and thereby enhances the function of PLP as an electron sink.

Correlation of polarized absorption spectroscopic and X-ray diffraction studies of crystalline cytosolic aspartate aminotransferase of pig hearts.

Absorption spectra of large, well-formed crystals of cytosolic aspartate aminotransferase have been recorded using plane polarized light. Making use of measurements of crystal thickness we have calculated extinction coefficients with the electric vector of the light parallel to both the a and c axes of the crystals of the enzyme in space group P2(1)2(1)2(1). The spectra have been resolved into components with lognormal distribution curves and the resulting integrated intensities have been used to calculate the c/a polarization ratios for the absorption bands of the bound co-enzyme pyridoxal 5'-phosphate. From the polarization ratio and the co-ordinates of the co-enzyme ring atoms, provided by X-ray crystallography, we have assigned principal molecular directions of the transition dipole moment within the plane of the co-enzyme ring. Of two possible orientations, only one predicts the correct crystal extinction coefficients for the 436 nm band. In this orientation, when viewed from the B face of the ring (i.e. looking into the active site of the enzyme), the transition moment is related to the N-1-C-4 axis of the ring by counterclockwise rotation by 27 degrees. A tentative assignment of the principal molecular directions of the transition moment has also been made for the 368 nm band of the high pH form of the enzyme. In each case, the plane of the co-enzyme ring was located from the atomic co-ordinates of the ring atoms and of those atoms attached directly to the ring. The projection of the N-1 to C-4 axis on to this plane was used to evaluate the orientation of the transition moment, which was presumed to lie precisely within the plane of the ring. We have tilted this plane systematically to evaluate the error in transition moment direction resulting from uncertainties in the atomic co-ordinates. When 2-methylaspartate is diffused into the crystals if forms a Schiff base with the co-enzyme in which the ring has tilted about 32 degrees from its original position and the polarization ratio of the 436 nm band drops from 1.6 in the free enzyme to about 0.38. On the assumption that the orientation of the transition moment within the co-enzyme does not change during this rotation, this value of the polarization ratio is within experimental error of that predicted from X-ray structures on the two forms. The 2-methylaspartate binds only to subunit 1.(ABSTRACT TRUNCATED AT 250 WORDS)

The effects of nested primer binding sites on the reproducibility of PCR: mathematical modeling and computer simulation studies.

The polymerase chain reaction (PCR) has become an indispensable tool in modern biological research. Although the application of PCR is a standard routine, we widely lack a theoretical understanding of the dynamic processes involved, especially with respect to the amplification of nonreproducible and/or unexpected amplification products. For one potential source of uncertainty, the presence of nested primer binding sites within an amplifyable DNA locus, we consider a simple stochastic model for the dynamics of PCR amplification of competing products. For commonly used thermostable DNA polymerases lacking a 5'-3'-exonuclease activity, we predict the relative amplification frequencies of competing PCR products dependent on the primer binding probability, the number of PCR cycles, and the number of initial DNA template molecules. At low primer binding probabilities and low numbers of initial DNA template molecules and PCR cycles, we expect the amplification of two products. At high primer binding probabilities and/or high copy numbers of initial template molecules only one main amplification product is predicted at increasing cycle numbers. Furthermore, by means of computer simulation studies we quantify the stochastic variation for the amplification frequencies of competing products.

Convulsant activity of pyridoxal sulphate and phosphonoethyl pyridoxal: antagonism by GABA and its synthetic analogues.

Pyridoxal phosphate and its synthetic analogues--pyridoxal 5'-sulphate and the 5-phosphonoethyl analogue of pyridoxal (phosphonoethyl pyridoxal) in doses of 0.125-0.250 (mumol/10 microliters/i.c.v./rat), caused epileptic seizures characterized by running fits, vocalization, muscular fasciculation and tonic-clonic convulsions. These effects were specific and could not be demonstrated with 5'-deoxypyridoxal, N-methylpyridoxal phosphate or the 5-trans-carboxyethenyl analogue of pyridoxal phosphate (carboxyethenyl pyridoxal). Structure-activity relationships of these analogues indicated that the presence of a CHO in position 4 of the pyridine ring was essential, since its conversion to CH2NH2 or CH2OH abolished activity. The presence of an unsubstituted N was essential, since convulsions did not occur with N-methylpyridoxal phosphate. The presence of the hydroxyl group in position 5' was essential since 5'-deoxypyridoxal was inactive. The convulsive activity was potentiated in the presence of both CHO and PO4, CHO and CH2--CH2PO2-4 but especially CHO and --OSO23-- groups. This seizure activity was prevented, attenuated or reversed by intracerebroventricular administration of 20 microliter of GABA (1 mumol), muscimol (0.025 mumol), trans-4-aminocrotonic acid (0.25 mumol), isoguvacine (0.25 mumol) or THIP (0.25 mumol), but not by biogenic amines. An understanding of the mechanism of pyridoxal phosphate-related seizures may provide additional insights not only about GABA receptor sites but also about the biochemical manifestation and expression of epilepsy.

Phosphonate analogues of pyridoxal phosphate with shortened side chains.

A phosphonate analogue of pyridoxal 5'-phosphate containing a 5'-phosphonomethyl group and its monoethyl and diethyl esters have been prepared. Except for the diethyl ester, the compounds appear to bind into the active site of aspartate aminotransferase. However, they lack detectable catalytic activity with this enzyme and with glutamate decarboxylase of Escherichia coli. The phosphonomethyl analogue bound to aspartate aminotransferase does react slowly with substrates, as determined by spectrophotometric observations; the monomethyl ester reacts about 20 times less rapidly. Because of the stability of the phosphonate linkage, these compounds may be useful as modifying reagents for various proteins.

Parenteral anticoagulation with the heparinoid Lomoparan (Org 10172) in patients with heparin induced thrombocytopenia and thrombosis.

Progressive thrombocytopenia may develop in as many as 5% of patients receiving heparin anticoagulation. In these patients, the risk of thromboembolic complications as well as continued thrombocytopenia necessitates discontinuation of heparin and initiation of an alternative anticoagulant when indicated. The heparinoid Lomoparan (Org 10172) is a mixture of several non-heparin low molecular weight glycosaminoglycans with proven anticoagulant efficacy that is generally non-reactive with platelets in the presence of plasma from patients with heparin induced thrombocytopenia, whereas standard heparin will induce platelet aggregation. We evaluated the role of heparinoid as a potential alternative anticoagulant in patients with heparin induced thrombocytopenia. During a 6 month period, we identified six patients with heparin induced thrombocytopenia who required an alternative parenteral anticoagulant, four as primary treatment for specific medical problem, and two as anticoagulation during a necessary surgical procedure. Heparinoid was used successfully in both medical and surgical patients requiring parenteral anticoagulation. In no case was there an exacerbation of the thrombocytopenia nor thromboembolic complications while on heparinoid therapy. Three of our patients sustained hemorrhagic complications, predominantly in the post-surgical setting in association with elevated anti-factor Xa levels and additional anticoagulant agents. We feel that these results confirm the utility of heparinoid anticoagulation in a select subset of patients with heparin induced thrombocytopenia who require continued parenteral anticoagulation.

Starch-binding domain of Aspergillus glucoamylase-I. Interaction with beta-cyclodextrin and maltoheptaose.

The characterization is reported of two peptide fragments (SBD106 and SBD122) containing the starch-binding domain (SBD) of Aspergillus sp. glucoamylase I. The starch-binding peptides were produced in Escherichia coli as fusion proteins of the maltose-binding protein (MBP). SBD106 (11.9 kDa) and SBD122 (13.8 kDa) were purified from the factor Xa digest of MBP fusion proteins. The amino acid compositions were similar to those deduced from their amino acid sequences. The interactions of beta-cyclodextrin and maltoheptaose with purified SBD peptides were investigated by UV difference spectroscopy. SBD106 and SBD122 bound specifically beta-cyclodextrin with a dissociation constant (Kd) of 34 microM and 23.5 microM, respectively. Maltoheptaose binding to SBD106 and SBD122 was weaker than that of beta-cyclodextrin; dissociation constants were 0.57 and 0.50 mM, respectively. The results indicate that the intramolecular disulfide bonding is not required for the domain functioning and that O-glycosylation is not critical for the functioning of the starch-binding domain, but may affect its conformation and dynamics.

Myo-inositol in depressive and healthy subjects determined by frontal 1H-magnetic resonance spectroscopy at 1.5 tesla.

Myo-inositol (mI) as a precursor in the phosphatidylinositol second messenger system has been reported to be reduced in depression. By means of proton-magnetic resonance spectroscopy (1H-MRS) the mI levels in the frontal brain were investigated in vivo in the present study. Twenty-two patients (mean age: 42.8 +/- 10.7 years) with depressive episodes according to ICD 10 (HAMD score > 17) were compared to 22 healthy subjects (28.0 +/- 5.3 years). Two voxels (30 x 20 x 20 mm3) in the frontal lobes were examined in a Siemens Magnetom SP 4000 at 1.5 T (STEAM sequence: TR = 3500 ms, TE = 55 ms). With the total creatine (Cr) as an internal standard, mI/Cr ratios were calculated to follow the mI levels. In the left frontal lobe, mI/Cr was 0.43 +/- 0.06 in depressive patients and 0.46 +/- 0.07 in healthy subjects; concerning the right frontal lobe, mI/Cr was 0.46 +/- 0.08 and 0.48 +/- 0.06, respectively. There were neither significant differences between the two groups nor between the hemispheres. Since there was a significant positive correlation (R = 0.6) between the age and the mI/Cr in the right frontal lobe of depressed patients, age matched pairs analysis was performed (n = 2 x 10, in each group: nine females, one male, < 40 years). In the right frontal lobe, the patients' mI/Cr of 0.40 +/- 0.05 was now significantly lower than the controls' mI/Cr of 0.45 +/- 0.06. However, most of the patients were on antidepressive medication. Interestingly, it was exactly this group of patients which showed significantly lower mI levels. We regard our investigation as a pilot study which suggests an influence of age and antidepressants on mI levels and should be taken into consideration in further investigations in depressive patients.

Patterns of antibiotic use and expenditures during 7 years at a university hospital.

Before 1982, annual retrospective drug-use audits clearly established the pattern of increased antibiotic use and expenditures at the University of Minnesota Hospital. At that point, a concurrent trend analysis system was developed to track this use by individual drug and therapeutic class, to identify trends in use for qualitative evaluation, and to provide an assessment of any intervention intended to alter use. Monthly use of all injectable antibiotics and antifungal agents, and selected oral antifungals has been tracked since 1982 as both defined daily doses and dollars. Use patterns in 1981 served as baseline. The defined daily dose index, which captures changes associated with intensity of antibiotic use per patient, increased 62% from baseline through 1988. This increase, however, resulted in a disproportionate 228% rise from baseline in antibiotic expenditures to nearly $3 million/year for 1988. The defined daily dose index peaked in 1985, declined in 1986, and remained stable for 1987 and 1988. Qualitative drug-use review programs were associated with this positive change. These results demonstrate the utility of concurrent trend analysis as a resource-management tool and as a measure of effectiveness for collaborative physician-pharmacist programs to foster safe, appropriate, and economical use of antiinfective agents.

Mental rotation: effects of dimensionality of objects and type of task.

The original studies of mental rotation estimated rates of imagining rotations that were much slower when two simultaneously portrayed three-dimensional shapes were to be compared (R. Shepard & J. Metzler) than when one two-dimensional shape was to be compared with a previously learned two-dimensional shape (Cooper and her associates). In a 2 X 2 design, we orthogonally varied dimensionality of objects and type of task. Both factors affected reaction times. Type of task was the primary determiner of estimated rate of mental rotation, which was about three times higher for the single-stimulus task. Dimensionality primarily affected an additive component of all reaction times, suggesting that more initial encoding is required for three-dimensional shapes. In the absence of a satisfactory way of controlling stimulus complexity, the results are at least consistent with the proposal that once three-dimensional objects have been encoded, their rotation can be imagined as rapidly as the rotation of two-dimensional shapes.

Absolute metabolite quantification by in vivo NMR spectroscopy: II. A multicentre trial of protocols for in vivo localised proton studies of human brain.

We have performed a multicentre trial to assess the performance of three techniques for absolute quantification of cerebral metabolites using in vivo proton nuclear magnetic resonance (NMR). The techniques included were 1) an internal water standard method, 2) an external standard method based on phantom replacement, and 3) a more sophisticated method incorporating elements of both the internal and external standard approaches, together with compartmental analysis of brain water. Only the internal water standard technique could be readily implemented at all participating sites and gave acceptable precision and interlaboratory reproducibility. This method was insensitive to many of the experimental factors affecting the performance of the alternative techniques, including effects related to loading, standing waves and B1 inhomogeneities; and practical issues of phantom positioning, user expertise and examination duration. However, the internal water standard method assumes a value for the concentration of NMR-visible water within the spectroscopic volume of interest. In general, it is necessary to modify this assumed concentration on the basis of the grey matter, white matter and cerebrospinal fluid (CSF) content of the volume, and the NMR-visible water content of the grey and white matter fractions. Combining data from 11 sites, the concentrations of the principal NMR-visible metabolites in the brains of healthy subjects (age range 20-35 years) determined using the internal water standard method were (mean+/-SD): [NAA]=10.0+/-3.4 mM (n=53), [tCho]=1.9+/-1.0 mM (n=51), [Cr + PCr]=6.5+/-3.7 mM (n=51). Evidence of system instability and other sources of error at some participating sites reinforces the need for rigorous quality assurance in quantitative spectroscopy.

Retention of multi-rooted teeth with class III furcation lesions utilizing resins. Report of 17 cases.

BACKGROUND: Treatment of periodontally diseased multi-rooted posterior teeth has conventionally consisted of scaling and root planing, apically positioned flaps, tunneling, root amputation, root resection, guided tissue regeneration, osseous grafting, and combination osseous graft and barrier membrane techniques. Successful treatment has often been directed and measured by how well the clinician obtains sufficient access to facilitate patient hygiene efforts. Long-term success is predicated upon tooth retention and the arrest of further destruction within the furcation area. METHODS: Seventeen adult periodontal patients with Class III furcation defects were evaluated. Using an open flap procedure, a resin-ionomer was placed into all 3 furcation defects. The patients were placed on quarterly maintenance appointments and the teeth evaluated up to 1 year. RESULTS: The results revealed that sealing the furcations of multi-rooted teeth with hopeless prognoses seems to be a viable alternative to accessing the same intraradicular areas for maintenance using more conventional modes of therapy. Sealing further decreases the surface area of the furca and simplifies future maintenance. CONCLUSIONS: The present study showed that teeth with hopeless prognoses might be retained by decreasing probing depths, bleeding upon probing, and mobility when furcation areas are sealed with a resin-ionomer.

Treatment of the palato-gingival groove with guided tissue regeneration. Report of 10 cases.

The purpose of this study is to demonstrate the potential of using a barrier in the treatment of palato-gingival groove defects. The study group consisted of 10 patients. Prior to treatment, the palato-gingival groove on maxillary lateral incisors was measured with calibrated periodontal probe from the cemento-enamel junction (CEJ) to the free gingival margin (FGM) and from the FGM to the base of the pocket (BP). Probing depth (PD) was calculated and bleeding on probing indicated. Surgical procedures consisted of flap reflection, removal of granulation tissue, and scaling and root planning of the groove. An expanded polytetrafluoroethylene membrane was sutured over the palato-gingival groove. Six months postsurgery, all measurements were repeated. Statistical analysis compared results using means, standard deviations, and paired t tests. Results showed an improvement in clinical attachment gain, probing depth reduction, and decreased bleeding on probing. This study demonstrates the potential of guided tissue regeneration in the treatment of palato-gingival groove defects. A random blinded clinical trial is necessary, however, to fully assess the potential of this procedure in treatment of palato-gingival groove defects.

Free gingival graft following biopsy: a case report of tissue management.

Excisional biopsies require wide margins to facilitate removal of diseased tissue. Such margins often result in soft-tissue defects. These defects can cause reduced esthetics, ineffective oral hygiene performance, and postsurgical root sensitivity. This case documents the combination of two dental procedures: biopsy and the free gingival graft. A pyogenic granuloma, as diagnosed by histologic report, was removed by excisional biopsy and the resultant defect repaired by using a free gingival graft. Palatal tissue was grafted immediately to the site of the biopsy and sutured in place to cover the loss of attached tissue and papilla. At 6 months, the graft appeared to restore gingival health and maintain both esthetics and function in the surgical biopsy site. The patient was well served by correcting the resultant biopsy defect in a one-step procedure which encouraged healing and an excellent esthetic result.