Soft Tissue Infection Model Research Articles

CcpA and LacD.1 affect temporal regulation of Streptococcus pyogenes virulence genes.

Production of H(2)O(2) follows a growth phase-dependent pattern that mimics that of many virulence factors of Streptococcus pyogenes. To gain greater insight into mechanisms coupling virulence factor expression to growth phase, we investigated the molecular basis for H(2)O(2) generation and its regulation. Deletion of the gene encoding lactate oxidase (lctO) or culture in the presence of glucose eliminated H(2)O(2) production, implicating carbohydrate regulation of lctO as a key element of growth phase control. In examining known carbohydrate-responsive regulators, deletion of the gene encoding CcpA but not that encoding LacD.1 resulted in both derepression and an uncoupling of lctO transcription from its growth phase pattern. Expanding this analysis to additional virulence factors demonstrated both negative (cfa, encoding CAMP factor) and positive (speB, encoding a cysteine protease) regulation by CcpA and that CcpA mutants were highly cytotoxic for cultured macrophages. This latter property resulted from enhanced transcription of the streptolysin S biogenesis operon. Examination of CcpA-promoter interactions using a DNA pull-down assay mimicking physiological conditions showed direct binding to the promoters of lctO and speB but not those of sagA. CcpA but not LacD.1 mutants were attenuated in a murine model of soft-tissue infection, and analysis of gene expression in infected tissue indicated that CcpA mutants had altered expression of lctO, cfa, and speB but not the indirectly regulated sagA gene. Taken together, these data show that CcpA regulates virulence genes via at least three distinct mechanisms and that disruption of growth phase regulation alters transcriptional patterns in infected tissues.

The Metal Homeostasis Protein, Lsp, of Streptococcus pyogenes Is Necessary for Acquisition of Zinc and Virulence

"Cluster 9" family lipoproteins function as ligand-binding subunits of ABC-type transporters in maintaining transition metal homeostasis and have been implicated in the virulence of several bacteria. While these proteins share high similarity, the specific metal that they recognize and whether their role in virulence directly involves metal homeostasis cannot be reliably predicted. We examined the cluster 9 protein Lsp of Streptococcus pyogenes and found that specific deletion of lsp produced mutants highly attenuated in a murine model of soft tissue infection. Under standard in vitro conditions, growth of the Lsp(-) mutant was indistinguishable from that of the wild type, but growth was defective under zinc-limited conditions. The growth defect could be complemented by plasmids expressing wild-type Lsp but not Lsp engineered to lack its putative lipidation residue. Furthermore, Zn(2+) but not Mn(2+) rescued Lsp(-) growth, implicating Zn(2+) as the physiological ligand for Lsp. Mutation of residues in the putative Zn(2+)-binding pocket generated variants both hypo- and hyper-resistant to zinc starvation, and both mutant classes displayed attenuated virulence. Together, these data suggest that Lsp is a ligand-binding component of an ABC-type zinc permease and that perturbation of zinc homeostasis inhibits the ability of S. pyogenes to cause disease in a zinc-limited host milieu.

Penicillin‐Binding Protein 7/8 Contributes to the Survival ofAcinetobacter baumanniiIn Vitro and In Vivo

Acinetobacter baumannii is a bacterial pathogen of increasing medical importance. Little is known about genes important for its survival in vivo. Screening of random transposon mutants of the model pathogen AB307-0294 identified the mutant AB307.27. AB307.27 contained its transposon insertion in pbpG, which encodes the putative low-molecular-mass penicillin-binding protein 7/8 (PBP-7/8). AB307.27 was significantly killed in ascites (P<.001), but its growth in Luria-Bertani broth was similar to that of its parent, AB307-0294 (P=.13). The survival of AB307.27 was significantly decreased in a rat soft-tissue infection model (P<.001) and a rat pneumonia model (P=.002), compared with AB307-0294. AB307.27 was significantly killed in 90% human serum in vitro, compared with AB307-0294 (P<.001). Electron microscopy demonstrated more coccobacillary forms of AB307.27, compared with AB307-0294, suggesting a possible modulation in the peptidoglycan, which may affect susceptibility to host defense factors. These findings demonstrate that PBP-7/8 contributes to the pathogenesis of A. baumannii. PBP-7/8 either directly or indirectly contributes to the resistance of AB307-0294 to complement-mediated bactericidal activity. An understanding of how PBP-7/8 contributes to serum resistance will lend insight into the role of this low-molecular-mass PBP whose function is poorly understood.

EXPERIMENTAL MODELS OF SEPSIS AND THEIR CLINICAL RELEVANCE

Sepsis remains a major cause of morbidity and mortality mainly because of sepsis-induced multiple organ dysfunction. In contrast to preclinical studies, most clinical trials of promising new treatment strategies for sepsis have failed to demonstrate efficacy. Although many reasons could account for this discrepancy, the misinterpretation of preclinical data obtained from experimental studies and especially the use of animal models that do not adequately mimic human sepsis may have been contributing factors. In this review, the potentials and limitations of various animal models of sepsis are discussed to clarify to which extent these findings are relevant to human sepsis. Such models include intravascular infusion of endotoxin or live bacteria, bacterial peritonitis, cecal ligation and perforation, soft tissue infection, pneumonia or meningitis models using different animal species including rats, mice, rabbits, dogs, pigs, sheep, and nonhuman primates. Despite several limitations, animal models remain essential in the development of all new therapies for sepsis and septic shock because they provide fundamental information about the pharmacokinetics, toxicity, and mechanism of drug action that cannot be replaced by other methods. New therapeutic agents should be studied in infection models, even after the initiation of the septic process. Furthermore, debility conditions need to be reproduced to avoid the exclusive use of healthy animals, which often do not represent the human septic patient.

Rat Pneumonia and Soft-Tissue Infection Models for the Study of Acinetobacter baumannii Biology

Acinetobacter baumannii is a bacterial pathogen of increasing medical importance. Little is known about its mechanisms of pathogenesis, and safe reliable agents with predictable activity against A. baumannii are presently nonexistent. The availability of relevant animal infection models will facilitate the study of Acinetobacter biology. In this report we tested the hypothesis that the rat pneumonia and soft-tissue infection models that our laboratory had previously used for studies of extraintestinal pathogenic Escherichia coli were clinically relevant for A. baumannii. Advantages of these models over previously described models were that the animals were not rendered neutropenic and they did not receive porcine mucin with bacterial challenge. Using the A. baumannii model pathogen 307-0294 as the challenge pathogen, the pneumonia model demonstrated all of the features of infection that are critical for a clinically relevant model: namely, bacterial growth/clearance, an ensuing host inflammatory response, acute lung injury, and, following progressive bacterial proliferation, death due to respiratory failure. We were also able to demonstrate growth of 307-0294 in the soft-tissue infection model. Next we tested the hypothesis that the soft-tissue infection model could be used to discriminate between the inherent differences in virulence of various A. baumannii clinical isolates. The ability of A. baumannii to grow and/or be cleared in this model was dependent on the challenge strain. We also hypothesized that complement is an important host factor in protecting against A. baumannii infection in vivo. In support of this hypothesis was the observation that the serum sensitivity of various A. baumannii clinical isolates in vitro roughly paralleled their growth/clearance in the soft-tissue infection model in vivo. Lastly we hypothesized that the soft-tissue infection model would serve as an efficient screening mechanism for identifying gene essentiality for drug discovery. Random mutants of 307-0294 were initially screened for lack of growth in human ascites in vitro. Selected mutants were subsequently used for challenge in the soft-tissue infection model to determine if the disrupted gene was essential for growth in vivo. Using this approach, we have been able to successfully identify a number of genes essential for the growth of 307-0294 in vivo. In summary, these models are clinically relevant and can be used to study the innate virulence of various Acinetobacter clinical isolates and to assess potential virulence factors, vaccine candidates, and drug targets in vivo and can be used for pharmacokinetic and chemotherapeutic investigations.

TRNA Modification by GidA/MnmE Is Necessary forStreptococcus pyogenesVirulence: a New Strategy To Make Live Attenuated Strains

Studies directed at vaccine development and mucosal immunity against Streptococcus pyogenes would benefit from the availability of live attenuated strains. Our approach for production of candidate live attenuated strains was to identify mutations that did not alter growth in vitro and did not alter the overall complement of virulence factors produced but did result in reduced levels of expression of multiple secreted virulence factors. A global reduction but not elimination of expression would likely lead to attenuation while maximizing the number of antigenic targets available for stimulation of immunity. Adaptation of Tn5-based transposome mutagenesis to S. pyogenes with initial screening for reduced expression of the SpeB protease resulted in identification of mutations in gidA, which encodes an enzyme involved in tRNA modification. Reduced SpeB expression was due to delayed onset of speB transcription resulting from reduced translation efficiency of the message for RopB, a transcriptional activator. Overall, GidA(-) mutants had a nearly normal global transcription profile but expressed significantly reduced levels of multiple virulence factors due to impaired translation efficiencies. A translation defect was supported by the observation that mutants lacking MnmE, which functions in the same tRNA modification pathway as GidA, phenocopied GidA deficiency. The mutants stimulated a cytokine response in cultured macrophages identical to that in the wild type, with the exception of reduced levels of tumor necrosis factor alpha and interleukin-23. Significantly, GidA(-) mutants were highly attenuated in the murine ulcer model of soft tissue infection. These characteristics suggest that GidA pathway tRNA modification mutants are attractive candidates for further evaluation as live attenuated strains.

A Chemokine-Degrading Extracellular Protease Made by Group AStreptococcusAlters Pathogenesis by Enhancing Evasion of the Innate Immune Response

Circumvention of the host innate immune response is critical for bacterial pathogens to infect and cause disease. Here we demonstrate that the group A Streptococcus (GAS; Streptococcus pyogenes) protease SpyCEP (S. pyogenes cell envelope protease) cleaves granulocyte chemotactic protein 2 (GCP-2) and growth-related oncogene alpha (GROalpha), two potent chemokines made abundantly in human tonsils. Cleavage of GCP-2 and GROalpha by SpyCEP abrogated their abilities to prime neutrophils for activation, detrimentally altering the innate immune response. SpyCEP expression is negatively regulated by the signal transduction system CovR/S. Purified recombinant CovR bound the spyCEP gene promoter region in vitro, indicating direct regulation. Immunoreactive SpyCEP protein was present in the culture supernatants of covR/S mutant GAS strains but not in supernatants from wild-type strains. However, wild-type GAS strains do express SpyCEP, where it is localized to the cell wall. Strain MGAS2221, an organism representative of the highly virulent and globally disseminated M1T1 GAS clone, differed significantly from its isogenic spyCEP mutant derivative strain in a mouse soft tissue infection model. Interestingly, and in contrast to previous studies, the isogenic mutant strain generated lesions of larger size than those formed following infection with the parent strain. The data indicate that SpyCEP contributes to GAS virulence in a strain- and disease-dependent manner.

RivR and the small RNA RivX: the missing links between the CovR regulatory cascade and the Mga regulon

The CovR/S two-component system regulates the transcription of many genes that are crucial for the virulence of Streptococcus pyogenes (group A Streptococcus, GAS). Previously, we demonstrated that one gene repressed directly by CovR is rivR, which encodes a member of the RofA-like family of transcriptional regulators. In this study, we deleted rivR and its downstream gene rivX in a DeltacovR background. Microarray analysis revealed that the products of the rivRX locus exert positive control over the transcription of members of the Mga regulon. Using mutational analysis, we established that rivX encodes a small regulatory RNA. We found that RivR enhances transcriptional activation by Mga in vivo and in vitro. An M1 DeltacovRDeltarivRX strain is attenuated for virulence in a murine model of invasive soft tissue infection and this attenuation is complemented by rivRX expressed from a plasmid, demonstrating the importance of the rivRX locus in pathogenesis. This study provides the first link between the CovR and Mga regulatory networks. By integrating the signals received through these two global regulators, GAS is able to select from its repertoire different combinations of specific virulence factors to express in response to a broad spectrum of environmental conditions.

Experimental models of sepsis and septic shock: an overview

Sepsis remains a major cause of morbidity and mortality in surgical patients and trauma victims, mainly due to sepsis-induced multiple organ dysfunction. In contrast to preclinical studies, most clinical trials of promising new treatment strategies for sepsis have fails to demonstrate efficacy. Although many reasons could account for this discrepancy, the misinterpretation of preclinical data obtained from experimental studies, and especially the use of animal models that do not adequately mimic human sepsis may have been contributing factors. In this review, the benefits and limitations of various animal models of sepsis are discussed to clarify the extend to which findings are relevant to human sepsis, particularly with respect to the subsequent design and execution of clinical trials. Such models include intravascular infusion of endotoxin or live bacteria, bacterial peritonitis, cecal ligation and perforation, soft tissue infection, pneumonia or meningitis models, using different animal species including rats, mice, rabbits, dogs, pigs, sheep and nonhuman primates. Despite several limitations, animal models remain essential in the development of all new therapies for sepsis and septic shock, because they provide fundamental information about the pharmacokinetics, toxicity, and mechanism of drug action that cannot be duplicated by other methods. New therapeutic agents should be studies in infection models, even after the initiation of the septic process. Furthermore, debility conditions need to be reproduced to avoid the exclusive use of healthy animals, which often do not represent the human septic patient.

A locus of group A Streptococcus involved in invasive disease and DNA transfer.

Group A streptococcus (GAS) causes diseases ranging from benign to severe infections such as necrotizing fasciitis (NF). The reasons for the differences in severity of streptococcal infections are unexplained. We developed the polymorphic-tag-lengths-transposon-mutagenesis (PTTM) method to identify virulence genes in vivo. We applied PTTM on an emm14 strain isolated from a patient with NF and screened for mutants of decreased virulence, using a mouse model of human soft-tissue infection. A mutant that survived in the skin but was attenuated in its ability to reach the spleen and to cause a lethal infection was identified. The transposon was inserted into a small open reading frame (ORF) in a locus termed sil, streptococcal invasion locus. sil contains at least five genes (silA-E) and is highly homologous to the quorum-sensing competence regulons of Streptococcus pneumoniae. silA and silB encode a putative two-component system whereas silD and silE encode two putative ABC transporters. silC is a small ORF of unknown function preceded by a combox promoter. Insertion and deletion mutants of sil had a diminished lethality in the animal model. Virulence of a deletion mutant of silC was restored when injected together with the avirulent emm14-deletion mutant, but not when these mutants were injected into opposite flanks of a mouse. DNA transfer between these mutants occurred in vivo but could not account for the complementation of virulence. DNA exchange between the emm14-deletion mutant and mutants of sil occurred also in vitro, at a frequency of approximately 10-8 for a single antibiotic marker. Whereas silC and silD mutants exchanged markers with the emm14 mutant, silB mutant did not. Thus, we identified a novel locus, which controls GAS spreading into deeper tissues and could be involved in DNA transfer.

Effects of coligand variation on the in vivo characteristics of Tc-99m-labeled interleukin-8 in detection of infection.

In our previous studies, interleukin-8 (IL-8) was labeled with (99m)Tc using hydrazinonicotinamide (HYNIC) as bifunctional coupling agent and tricine as coligand. This preparation showed excellent characteristics for imaging of infection in a rabbit model of soft-tissue infection. In the present study, the propylaldehyde hydrazone formulation of HYNIC was introduced to stabilize HYNIC-IL-8. (99m)Tc-HYNIC-IL-8 was prepared using 5 different coligand formulations. The effect of these coligand formulations on the in vitro characteristics and in vivo behavior of (99m)Tc-HYNIC-IL-8 was investigated. HYNIC-conjugated IL-8 was labeled with (99m)Tc in the presence of either (A) tricine, (B) ethylenediaminediacetic acid (EDDA), (C) tricine and trisodium triphenylphosphinetrisulfonate (TPPTS), (D) tricine and nicotinic acid (NIC), or (E) tricine and isonicotinic acid (ISONIC). These preparations were characterized in vitro by RP-HPLC, determination of the octanol/water partition coefficient, stability studies, and receptor binding assays. The in vivo biodistribution of the radiolabel in rabbits with E. coli-induced soft-tissue infection was determined both by gamma-camera imaging as well as by tissue counting at 6 h pi. Specific activity (MBq/microg) was highest for (ISO)NIC (up to 80) > TPPTS (40) > tricine (15) > EDDA (7). RP-HPLC and octanol/water partition coefficients showed a shift toward higher lipophilicity for the TPPTS preparation. The leukocyte receptor binding fractions were around 40-55% for all preparations except for TPPTS, which showed predominantly nonspecific binding. All preparations were stabilized in serum, but the stability in PBS was highest for NIC and TPPTS > EDDA > ISONIC > tricine. The in vivo biodistribution showed highest abscess/muscle for NIC and ISONIC (>200) > EDDA and tricine (approximately 100) > TPPTS (<40). Gamma camera imaging rapidly visualized the abscess from 2 h pi onward for all formulations. The abscess/background (A/B) at 6 h pi for ISONIC was significantly higher (P < 0.05) than that of tricine and the A/B of TPPTS was significantly lower (P < 0.05). IL-8 can be rapidly and easily labeled with (99m)Tc using HYNIC as a chelator in combination with various coligands. The most optimal infection imaging characteristics were found for formulations using nicotinic acid/tricine as coligand system combining a high specific activity and high in vitro stability with high abscess/muscle ratios (>200) and high abscess/background ratios (>20). Protein doses to be administered were as low as 70 ng/kg bodyweight. At these low protein doses, side effects are not to be expected in the human system. This paves the way for infection imaging studies in patients.

Antimicrobial and clinical effect of linezolid (ZYVOX), new class of synthetic antibacterial drug

Linezolid (ZYVOX), a novel synthesized antibacterial drug, was first approved in April 2001, as an antibacterial against vancomycin (VCM)--resistant enterococci in Japan. LZD has a wide spectrum of antibacterial activity against gram-positive bacteria with MIC90 of 0.5-4 mcg/mL. These antibacterial activities of LZD are similar to those of vancomycin (VCM). LZD also has similar antibacterial activities against drug-resistant bacteria including VRE and MRSA. Protein-synthesis inhibitors, e.g., macrolides, tetracycline, aminoglycosides, and chloramphenicol, are known to bind the 30S and 50S subunits of ribosomes and inhibit the elongation cycle of protein synthesis. In contrast, LZD was found to inhibit the process of formation of the 50S, 30S-mRNA, and fMet-tRNA complex in the ribosome cycle, but not the elongation cycle. Due to this novel mechanism of action, LZD does not have a cross-resistance to drug-resistant bacteria and development of its resistance is quite slow. The antibacterial activity of LZD against VRE is bacteriostatic. In vivo antibacterial activity of orally administered LZD was demonstrated in a mouse model of systemic infection by VRE. When administered orally prior to the abscess formation in a mouse model of soft tissue infection by VRE, LZD showed similar antibacterial activity against VRE infection to that against VCM-susceptible enterococci. LZD is rapidly absorbed following oral administration and bioavailability when compared with intravenous administration is almost 100%. LZD administered orally twice-daily showed excellent efficacy in clinical trials with VRE-infected patients.

Subacute Sepsis Impairs Vascular Smooth Muscle Contractile Machinery and Alters Vasoconstrictor and Dilator Mechanisms

Introduction.Sepsis results in hyporesponsiveness to α-adrenergic stimulation. This is thought to be mediated by the release of vasoactive compounds from the septic endothelium or by the direct effect of sepsis on vascular smooth muscle (VSM) contractile mechanics and machinery. Previous studies have used lethal models of sepsis or endotoxemia to examine this phenomenon. The present study utilizes a clinically relevant, nonlethal model of soft tissue infection to determine the effects of sepsis on α-adrenergic mechanisms. We hypothesize that subacute sepsis causes impaired α-adrenergic vascular responsiveness by a combination of effects on adrenergic constrictor mechanisms, endogenous dilator tone, and VSM contractile function.Methods.Male Sprague-Dawley rats underwent implantation of a 2 × 2-cm2gauze sponge into a subcutaneous pocket created at the base of the tail. Five days after implantation, sepsis (S) was induced by inoculation of the sponge with 109CFUEscherichia coliandBacteroides fragilis.Controls (C) were inoculated with saline. Thoracic aortic harvest was performed 24 and 48 h after sponge inoculation for organ bath ring studies. Receptor-mediated (phenylephrine) and nonreceptor-mediated (KCl) maximum force of contraction (Fmax) was measured. Vessel sensitivity (pD2) to phenylephrine, acetylcholine, and KCl was calculated from dose-response curves.Results.At 24 h, sepsis resulted in a lowerFmaxto phenylephrine (1.15 for C vs 0.5 for S,P< 0.05 by ANOVA), despite an increase in vessel sensitivity (pD2) to α-adrenergic stimulation (6.70 for C vs 6.88 for S,P< 0.05 by ANOVA).Fmaxto KCl was lower in septic animals at 24 h (3.50 for C vs 2.77 for S,P< 0.05 by ANOVA) and sensitivity to acetylcholine (pD2) was markedly increased (6.56 for C vs 7.23 for S,P< 0.05 by ANOVA). At 48 h, the impairment inFmaxto α-adrenergic stimulation (2.29 for C vs 1.72 for S,P< 0.05 by ANOVA) and KCl (3.5 for C vs 3.08 for S.P< 0.05 vs 24 h C by ANOVA) persisted without any change in sensitivity to phenylephrine or acetylcholine.Conclusions.Subacute sepsis results in an early suppression of maximum contractile force despite an increase in adrenergic receptor sensitivity (pD2). This may be secondary to an elevation in dilator sensitivity combined with a direct effect of sepsis on VSM contractile mechanisms. Later in the septic process, however, α-adrenergic hyporesponsiveness (↓Fmax) is primarily due to changes in VSM contractile machinery.

In vivo activities of U-100592 and U-100766, novel oxazolidinone antimicrobial agents, against experimental bacterial infections

The Upjohn oxazolidinones, U-100592 and U-100766, are orally bioavailable synthetic antimicrobial agents with spectra of activity against antibiotic-susceptible and -resistant gram-positive pathogens. In several mouse models of methicillin-resistant Staphylococcus aureus infection, U-100592 and U-100766 yielded oral 50% effective doses (ED50) ranging from 1.9 to 8.0 mg/kg of body weight, which compared favorably with vancomycin subcutaneous ED50 values of 1.1 to 4.4 mg/kg. Similarly, both compounds were active versus a Staphylococcus epidermidis experimental systemic infection. U-100592 and U-100766 effectively cured an Enterococcus faecalis systemic infection, with ED50 values of 1.3 and 10.0 mg/kg, and versus a vancomycin-resistant Enterococcus faecium infection in immunocompromised mice, both drugs effected cures at 12.5 and 24.0 mg/kg. Both compounds were exceptionally active in vivo against penicillin- and cephalosporin-resistant Streptococcus pneumoniae, with ED50 values ranging from 1.2 to 11.7 mg/kg in systemic infection models. In soft tissue infection models with S. aureus and E. faecalis, both compounds exhibited acceptable curative activities in the range of 11.0 to 39.0 mg/kg. U-100766 was also very active versus the Bacteroides fragilis soft tissue infection model (ED50 = 46.3 mg/kg). In combination-therapy studies, both U-100592 and U-100766 were indifferent or additive in vivo against a monomicrobic S. aureus infection in combination with other antibiotics active against gram-positive bacteria and combined as readily as vancomycin with gentamicin in the treatment of a polymicrobic S. aureus-Escherichia coli infection. U-100592 and U-100766 are potent oxazolidinones active against antibiotic-susceptible and -resistant gram-positive pathogens in experimental systemic and soft tissue infections.

Response of Pasteurella haemolytica to erythromycin and dexamethasone in calves with established infection

Summary A subcutaneous soft tissue infection model in calves was used to study the in vivo response of Pasteurella haemolytica to erythromycin and dexamethasone. Two tissue chambers were implanted sc in each of 12 calves. At 45 days after implantation, all tissue chambers were inoculated with an erythromycin-sensitive strain of P haemolytica. Starting 24 hours after inoculation, calves were allotted to 4 groups of equal size and a 2 × 2-factorial arrangement of treatments was applied: 3 calves were given erythromycin (30 mg/kg of body weight, im, for 5 days), 3 calves were given dexamethasone(0.05 mg/kg, im, for 2 days), 3 calves were given erythromycin and dexamethasone, and the remaining calves served as nontreated controls. Chamber fluids were tested daily, and the response to treatment was measured. Neither erythromycin nor dexamethasone affected viability or growth of bacteria within tissue chambers. Dexamethasone had no effect on the influx of neutrophils into infected chambers. Despite repeated administration of a high dose of erythromycin and attainment of adequate concentration in serum, erythromycin concentration in chamber fluids did not exceed the minimal inhibitory concentration established in vitro. These results indicate that the clinical efficacy of erythromycin against P haemolytica sequestered in consolidated pneumonic lesions may not be well correlated with predictions based on serum pharmacokinetic and in vitro susceptibility data.

The Effect of Venous Obstruction in Infected Pedicle Flap

A new model of soft-tissue infection is used to investigate the effect of the local wound environment on the septic focus. Island pedicle flaps were raised on the buttock of 24 adult ewes and multiply inoculated with Staphylococcus aureus. Flaps with bacterial inoculation, without compromise of venous outflow, showed distal necrosis (mean +/- SEM percent of surface area, 25.8% +/- 8.6%) and developed septic foci with bacterial counts one log less than the amount injected. Flaps with inoculation and venous outflow obstruction underwent subtotal necrosis (mean percent of surface area, 73.3% +/- 11.2%) and had counts two logs higher than the nonobstructed flaps but without discrete septic foci. Flaps without inoculation, with or without venous obstruction, survived completely. Venous outflow obstruction is shown herein to potentiate tissue necrosis by raising bacterial counts in a septic focus and preventing defensive abscess formation by the host.

Imaging infections with antibodies: A quantitative autoradiographic analysis

Radiolabeled IgG has recently been demonstrated to effectively image infections. A potential but unproven mechanism for this localization is the specific binding of IgG to Fc receptors on the surface of inflammatory cells in infections. In an animal model of soft tissue infection, quantitative autoradiography was used to measure 125I-labeled IgG and albumin in tissues with a spatial resolution sufficient to associate these proteins with cellular morphology. Gamma camera images at 24 h localized the infection with target-to-background ratios of 2.2 ± 0.5 for IgG and 2.3 ± 1.0 for albumin (mean ± SD). Using quantitative autoradiography at 1 h post-injection, significantly higher concentrations were found in infected thighs of 2–4% of initial plasma concentrations ( C Po) as compared to 0.2–0.3% of C Po in noninfected thighs ( P < 0.05); at 24 h post-injection, higher concentrations (7–8% of C Po) were found in infected thighs. Radiolabeled proteins were not inflammatory cell associated and were localized primarily within the edematous interstital spaces of the infection.

Characterization of a soft-tissue infection model in the horse and its response to intravenous cephapirin administration.

A soft-tissue infection model was created in eight horses by infecting subcutaneous tissue chambers with Streptococcus zooepidemicus organisms. Responses of the horses to the infections were determined by monitoring changes in the complete blood count and body temperature and by following changes in the cytology and protein content of the tissue chambers. Systemic reactions to the infections included a mild neutrophilia, mild pyrexia and mild anemia. There was a marked influx of neutrophils and protein into the chambers after they were seeded with bacteria and chamber neutrophil viability decreased markedly at the height of the infection. Subsequent to establishing tissue chamber infections four of the horses were treated with intravenous cephapirin t.d. at a dosage of 20 mg/kg for 5 days. Quantitative culturing of tissue chamber fluid was performed to analyze the efficacy of cephapirin therapy. Cephapirin therapy was accompanied by decreases in the systemic neutrophilia, pyrexia, anemia, and chamber bacterial counts. However, cephapirin did not eliminate the infection in any of the chambers. Chamber neutrophil viability was markedly increased during the cephapirin therapy period.