The erythromycin-resistance in S. pyogenes does not limit the human polymorphonuclear cell antimicrobial activity.

In order to highlight the potential erythromycin immunomodulatory properties related to different antibiotic resistance patterns in Streptococcus spp., we evaluated the influence of the macrolide on the PMNs primary functions against erythromycin-susceptible (Ery-S) and erythromycin-resistant (Ery-R) S. pyogenes strains. A total of 438 S. pyogenes were isolated over the period 2005-2007. On the basis of the triple disk testing, 345 out of 438 S. pyogenes isolates were Ery-S and 93 were Ery-R; among the resistant strains, 65 displayed the cMLSB phenotype, 23 had the M phenotype and 5 had iMLSB phenotype. Concerning antibacterial activity of PMNs, our results showed that erythromycin did not modify bacterial uptake, but significantly increased the phagocyte intracellular killing, compared with controls, for both Ery-S and Ery-R strains. Consequently, this report underlines that in immunocompetent hosts the dichotomy between the in vitro resistance and clinical trial data for antimicrobial agents should be thoroughly re-evaluated.

Role of fosfomycin tromethamine in modulating non-specific defence mechanisms in chronic uremic patients towards ESBL-producing Escherichia coli.

Antimicrobial agents and polymorphonuclear cells (PMNs) have the potential to interact in such a way that improve the therapy for infectious diseases. In immunocompromised patients highly susceptible to microbial infections with high morbidity and mortality, several metabolic and functional alterations in PMNs, mostly related to microbicidal activity, are observed. Therefore, the antibiotic of choice should have a good antimicrobial effect without impairing host defences. The aim of this study is to evaluate in vitro effects of sub-inhibiting fosfomycin tromethamine (FT) concentrations on the primary functions of PMNs from healthy subjects and immunocompromised patients (haemodialysed and renal transplant recipients), against an ESBL-producing Escherichia coli, the most common aetiological agent in urinary tract infections (UTIs). FT is considered a first line drug in the eradication of UTIs due to its appropriate antimicrobial spectrum, oral bioavailability and minimal risk of microbial resistance. Our results provide evidence that FT is able to induce enhancement of the depressed phagocytic response of PMNs from patients on chronic haemodialysis and from renal transplant recipients, restoring their primary functions in vitro against ESBL-producing E. coli. All these data permit the conclusion that uremic-infected patients might additionally benefit from the immunomodulating properties of FT.

Improvement of clinical response in allergic rhinitis patients treated with an oral immunostimulating bacterial lysate: in vivo immunological effects.

Allergic rhinitis is known to be one of the most common chronic diseases in the industrialized world. According to the concept that allergic rhinitis patients generally suffer from an immune deficit, in order to stimulate specifically or aspecifically their immune system, immunomodulating agents from various sources, such as synthetic compounds, tissue extracts or a mixture of bacterial extracts, have been used. The aim of the present trial is to evaluate the efficacy of the treatment with an immunostimulating vaccine consisting of a polyvalent mechanical bacterial lysate (PMBL) in the prophylaxis of allergic rhinitis and subsequently to analyze its in vivo effects on immune responses. 41 allergic rhinitis patients were enrolled: 26 patients were randomly assigned to the group for PMBL sublingual treatment and 15 others to the group for placebo treatment. For all 26 patients blood samples were drawn just before (T0) and after 3 months of PMBL treatment (T3) to evaluate plasma IgE levels (total and allergen-specific) and the cytokine production involved in the allergic response (IL-4, IFN-gamma). The results of our study indicate that PMBL is effective in vivo in the reduction or in the elimination of the symptoms in rhinitis subjects during the treatment period in comparison to a non-immunostimulating treatment. A significant and clinically relevant improvement was found in 61.5%, a stationary clinical response was registered in 38.4% and no negative side effects associated with the medication or worsening were recorded. At the end of a 3-month follow up period the clinical picture remained the same as that observed at T3. PMBL treatment did not affect the serum IgE levels (either total or allergen-specific) and did not induce significant changes in IFN-gamma concentration. In contrast, PMBL therapy may be accompanied, in some patients, by a potential immunomodulating activity by decreasing IL-4 cytokine expression.

Improved phagocyte response by co-amoxiclav in renal transplant recipients.

BACKGROUND: Infectious diseases are a major source of morbidity and mortality for immunosuppressed transplant recipients and the antimicrobial chemotherapy can be often less effective in these individuals, because the contribution of underlying host defenses is absent. METHODS: The influence of co-amoxiclav on the functions of polymorphonuclear granulocytes (PMNs) from renal transplant recipients were investigated. RESULTS: PMNs from renal transplant recipients showed a diminished phagocytic activity with reduced phagocytosis and bactericidal activity against intracellular Klebsiella pneumoniae, compared to that seen with PMNs from healthy subjects. Co-amoxiclav significantly elicited the functions of PMNs from uremic patients, resulting in an increased percentage of ingested klebsiellae and in a higher bactericidal effect (98-99%), compared with the drug-free control system. When PMNs were collected from renal transplant recipients treated with co-amoxiclav a significant high increase in both phagocytosis and killing activity were detected, showing the co-amoxiclav capability of "restoring" even in vivo the depressed primary functions of PMNs. CONCLUSIONS: The interesting beneficial properties of co-amoxiclav, which result in restoring the phagocyte-dependent response in renal transplant patients both in vitro and in vivo, may make this drug more suitable for the treatment of infections in patients with defects of phagocyte functions.

Impact of co-amoxiclav on polymorphonuclear granulocytes from chronic hemodialysis patients.

Phagocyte-dependent host defenses are frequently impaired in maintenance hemodialysis patients who show an increased susceptibility to infections. In these individuals, the course of infections can be more aggressive than in normal hosts, and the antibiotic of choice should have a high antimicrobial effect without impairing host defenses. Hence, in uremic patients, the antibiotic enhancement of phagocyte functions may be of potential clinical importance in the outcome of bacterial infections. Because we demonstrated previously that co-amoxiclav had beneficial properties that result in enhancement of the microbicidal functions of human polymorphonuclear cells (PMNs) from healthy subjects, we investigated the influence of this combination on the activities of PMNs from chronic hemodialysis patients against Klebsiella pneumoniae, a human pathogen that can pose severe problems in patients whose immunity is impaired. PMNs from chronic dialysis patients showed a diminished in vitro phagocytic efficiency with a reduced phagocytosis and bactericidal activity towards intracellular K. pneumoniae compared with that seen in PMNs from healthy subjects. When co-amoxiclav was added to PMNs from chronic hemodialysis patients, it was able to restore the depressed primary functions of PMNs, resulting in a significant high increase in both phagocytosis or killing activity. A similar pattern was detected with PMNs collected from hemodialysis patients treated with co-amoxiclav. The results of the present study provide evidence that co-amoxiclav is able to induce stimulation of depressed phagocytic response of PMNs from patients on chronic hemodialysis, restoring their primary functions both in vitro and in vivo.

Effect of bacampicillin and ampicillin on the phagocytic, microbicidal functions of human macrophages.

During acute bacterial infections, antibiotics only assist the phagocytic system in eradicating the invading organism. In this regard, the effect of ampicillin and its prodrug bacampicillin on the phagocytosis and killing of Staphylococcus aureus by human macrophages was compared. Bacterial phagocytosis was enhanced after pre-incubation of staphylococci with bacampicillin. At a concentration of half the MIC bacampicillin showed a much more impressive killing capacity on macrophage-ingested live bacteria than did ampicillin, under all the experimental conditions tested.

Teicoplanin and its influence on Staphylococcus aureus ultrastructure.

Teicoplanin, a glycopeptide antibiotic chemically related to the vancomycin-ristocetin group of antibiotics, has been shown to be active against gram-positive bacteria. Binding of teicoplanin by actively growing bacteria leads to morphological damage to the cell surface in staphylococci, as largely shown by transmission and scanning microscopy results.

New imidazole derivates: in-vitro activity on dermatophytes.

The in-vitro activities of two new miconazole and econazole salts (sulfosalicylic acid formulation) against 98 clinical isolates of dermatophytes were evaluated, in comparison to those of miconazole, miconazole nitrate, econazole and econazole nitrate. Miconazole sulfosalicylate and econazole sulfosalicylate exhibited good activity towards all the dermatophytes tested, although econazole and its derivatives were more efficacious than miconazole and its salts. The new imidazoles were equally effective in inhibiting the fungal growth with respect to miconazole, miconazole nitrate, econazole and econazole nitrate.

Interactions of antibiotics with phagocytes in vitro.

The goal of antimicrobial therapy is to eliminate invading bacteria both in the extracellular and intracellular environment. Thus, recent work has focused on the ability of various antibiotics to enter phagocytic cells and kill intracellular pathogens, since bacteria which survive within phagocytes may often produce prolonged or recurrent infections. In the last ten years the antibiotic modulation of phagocytic cell functions has become the subject of our major investigational activity. Taking into account that entry of antibiotics into phagocytes is necessary for activity against intracellular organisms, we examined the uptake of 11 radiolabelled antibiotics by macrophages. Penicillins and cephalosporins were taken up poorly by phagocytes. Teicoplanin was efficiently concentrated by macrophages, achieving intracellular concentrations higher than those in the surrounding extracellular medium. Roxithromycin was more markedly accumulated within phagocytes than was erythromycin, depending upon an active transport mechanism. Antitubercular drugs were concentrated approximately twofold in the macrophages. Besides, since antibiotics that act on phagocytosed bacteria have clinical advantages, we evaluated the direct action of the above mentioned drugs on the macrophage, by determining their interference with its functions or their potentiation of both phagocytosis and killing of bacteria.

Effect of ceftriaxone on the phagocytosis and intracellular killing of Staphylococcus aureus by human macrophages.

The efficacy of an antimicrobial agent in the treatment of infections depends upon the interactions of bacteria, antibiotic and phagocytes. The influence of ceftriaxone on the phagocytic and bactericidal activity of human macrophages in vitro and ex vivo was investigated. At concentrations one-half the minimum inhibitory concentration (1/2 x MIC) the antibiotic caused in vitro a significant enhancement of phagocytosis and a reduction in the survival of intracellular Staphylococcus aureus. The distinction between any effect of ceftriaxone on the staphylococci and the macrophages was made by exposure of each of them to the antibiotic before they were incubated together. The results suggest that ceftriaxone may have a direct positive action on macrophages, possibly by interfering with the cellular membrane functions and hence enhancing engulfment of bacteria. The ex vivo data seem to corroborate this hypothesis.

Interaction of Candida albicans, macrophages and fluconazole: in vitro and ex vivo observations.

In recent years, medical interest in evaluating the interaction among mycetes, phagocytes, and antimycotic drugs has increased notably due to higher incidences of fungal infections in immunocompromised subjects and to the long-term therapy they require. In this study the in vitro and ex vivo interaction of fluconazole, at plasma concentrations, with mouse macrophages was evaluated in the presence of different inocula of Candida albicans. The results showed that fluconazole did not interfere negatively with phagocyte functions; conversely, according to different experimental conditions, it was able to increase both phagocytosis and intracellular killing of candida, probably exerting its action on the yeast rather than on the phagocyte. A higher enhancement of macrophage functions was observed in vitro when the drug was present in the medium with macrophages and candida in a 1:1 ratio.

Susceptibility of clinical isolates of gram-positive and gram-negative organisms to cefatrizine.

The in vitro activity of cefatrizine was evaluated against 294 Gram-positive and 270 Gram-negative bacteria isolated from clinical specimens. Cefatrizine had excellent activity against Gram-positive cocci, inhibiting all except enterococci at minimal inhibitory concentrations below 1 mcg/ml. Moreover, cefatrizine was an effective antibacterial agent for most major Gram-negative species. Cefatrizine activity was tested against 34 strains of H. influenzae and 37 strains of K. pneumoniae and compared with that of other orally administered cephalosporins. Cefatrizine MIC50 values were much lower than those recorded for cephalexin and cefadroxil.

Imipenem: morphological changes and lethal effects on Pseudomonas aeruginosa.

Imipenem, a new carbapenem antibiotic shows an extremely broad spectrum of antibacterial activity for almost all Gram-negative and Gram-positive aerobic and anaerobic bacteria. It is stable to beta-lactamases and shows a high affinity for PBP 2. The changes in morphology and ultrastructure caused by the antibiotic on Ps. aeruginosa confirm that imipenem acts by binding primarily to PBP 2, resulting in irregular and round shaped cells, and later during treatment to PBP 1 with cellular lysis. The involvement of PBP 1 is also demonstrated by the fast bactericidal kinetics on Ps. aeruginosa, E. coli and Staph. aureus.

Potentiation of human polymorphonuclear leukocyte phagocytosis and intracellular bactericidal activity by amoxycillin/clavulanic acid.

The use of broad spectrum beta-lactamase inhibitors in association with beta-lactam agents provides one strategy to overcome the enzymatic resistance. Clavulanic acid is a potent inhibitor of a wide range of bacterial beta-lactamases and its potentiating effect on amoxycillin has been established both in vitro and in clinical trials. Since the efficacy of an antimicrobial agent in the therapy of infections depends on the interaction of bacteria, antibiotic and phagocytes, we investigated the effect of amoxycillin/clavulanic acid on the in vitro interaction between human polymorphonuclear cells (PMNs) and beta-lactamase producing strains of Klebsiella pneumoniae and Staphylococcus aureus. Clavulanic acid did not have any significant influence upon the PMN phagocytosis and killing against intracellular bacteria. Interestingly, the presence of the suicide inhibitor, with its beta-lactamase inhibitory properties, potentiated the activity of amoxycillin against the beta-lactamase producing strains of K. pneumoniae and S. aureus in such a manner that bacteria became significantly more susceptible to either phagocytosis or microbicidal activities of human phagocytes, compared to both the control and amoxycillin systems.

Cellular uptake and intraphagocytic activity of the new fluoroquinolone AF 3013 against Klebsiella pneumoniae.

The entry of an antibiotic into phagocytes is a prerequisite for its intracellular bioactivity against susceptible facultative or obligatory intracellular microorganisms. AF 3013 is a new fluoroquinolone, and its uptake into and elimination from mouse peritoneal macrophages, together with its effects on phagocytic and antimicrobial mechanisms against Klebsiella pneumoniae, were investigated. AF 3013 efficiently penetrated into phagocytic cells at all concentrations tested. The uptake proceeded rapidly and was energy independent, since it was not affected by cell viability, environmental temperature or the addition of a metabolic inhibitor. Therefore, a possible passive transmembrane diffusion mechanism might be proposed. The elution of AF 3013 from macrophages occurred relatively slowly; in fact, 60 min after the removal of extracellular AF 3013, nearly 40% of the drug still remained in the phagocytes. Exposure to 1 MIC of AF 3013 significantly enhanced macrophages phagocytosis and increased intracellular bactericidal activity against K. pneumoniae. Following preexposure of macrophages to 1 MIC of AF 3013, there was a significant increase in both phagocytosis and killing, compared with the controls, indicating the ability of AF 3013 to interact with biological membranes and remain active within phagocytes. Preexposure of Klebsiella to AF 3013 made the bacteria more susceptible to the bactericidal mechanisms of macrophages than untreated organisms.

Enhanced effects of amoxycillin/clavulanic acid compared with amoxycillin and clavulanic acid alone on the susceptibility to immunodefenses of a penicillin-resistant strain of Streptococcus pneumoniae.

The recent increase in the incidence of infections due to Streptococcus pneumoniae resistant to penicillin and other antibiotics, often associated with considerable morbidity and mortality, has been recognized as an alarming problem. From the recent medical literature data it emerges that among beta-lactam antibiotics used as an empiric treatment for infections caused by S. pneumoniae, amoxycillin and amoxycillin/clavulanic acid are the most active oral antibiotics and may be considered as a first-line therapeutic agent for the treatment of these infections. Since the therapeutic result of the treatment of an infection is determined by the combined effect of the antimicrobials and host defenses, we investigated the effect of amoxycillin, with and without clavulanic acid, upon the in vitro interaction between human polymorphonuclear leukocytes (PMNs) and a penicillin-resistant strain of S. pneumoniae. Amoxycillin significantly inhibited the streptococcal uptake by PMNs referred to the control system. Clavulanic acid did not have any significant effect upon the interaction PMNs-S. pneumoniae. The addition of amoxycillin/clavulanic acid to phagocytes and streptococci resulted in a synergystic potentiation of the activity of both drugs upon the PMN functions towards S. pneumoniae in such a manner that the bacteria became more susceptible to either the phagocytosis or the microbicidal activities of phagocytes. These effects came in addition to the intrinsic, excellent antimicrobial properties of this drug combination. Although the clinical significance of the observed enhanced effects of amoxycillin/clavulanate are far from elucidated, the possibility exists that they may play a contributory role, especially in patients with impaired host defense.

Phagocytosis and killing of cefazolin-exposed Staphylococci and Bacilli by mouse peritoneal macrophages.

The phagocytic and microbicidal capacity of proteose peptone-induced macrophages from Balb/c mice was assessed in vitro against Staphylococcus aureus and Bacillus subtilis in the presence of a subinhibitory concentration of cefazolin. Enhanced phagocytosis (50-60%) and a similar killing effect (99%) after two hours, incubation was noted towards both test organisms. Electron microscopy showed phagocytic activity on both control and antibiotic-treated strains. Since cefazolin does not penetrate macrophages, the peritoneal cell morphology and ultrastructure were unaffected by the drug.

Relationship between growth rate and phagocytosis susceptibility in Streptococcus mutans.

Streptococcus mutans strain GS-5 was grown in a variety of carbon sources in order to achieve different balanced growth rates, ranging from 61 to 383 minutes. The influence of the S. mutans growth rate on mouse phagocyte activity against these bacteria has been evaluated. The percentages of bacteria phagocytized and intracellularly killed by macrophages rose to 60-80% and 85-95% respectively when the doubling time was longer, showing that S. mutans is particularly sensitive to nonspecific immune defence mechanisms when cultured under conditions similar to those of its natural ecosystem.

Pefloxacin and immunity: cellular uptake, potentiation of macrophage phagocytosis and intracellular bioactivity for Klebsiella pneumoniae.

Antibiotic potentiation of host defence mechanisms may be of potential clinical importance in the outcome of infections. Therefore the effect of pefloxacin upon the interaction of in vitro of human macrophages with Klebsiella pneumoniae, by assays of antibiotic cellular uptake, bacterial phagocytosis and intracellular killing, was examined. The results indicated that pefloxacin was well concentrated by phagocytes at all the concentrations tested. The uptake proceeded rapidly and was not affected either by cell viability or physiological environmental temperature. Synergistic phagocytosis and intracellular killing of K. pneumoniae was observed in the presence of macrophages and subinhibitory concentrations (one-half MIC) of pefloxacin. Pretreatment of bacteria with pefloxacin led to an increase in both bacterial uptake and microbicidal activity of phagocytes. Exposure of the macrophages to pefloxacin did not affect any phagocyte functions.

Clarithromycin mediated the expression of polymorphonuclear granulocyte response against streptococcus pneumoniae strains with different patterns of susceptibility and resistance to penicillin and clarithromycin.

There is an urgent need for antibiotics that can be used in the therapy of infections caused by penicillin-resistant Streptococcus pneumoniae, the incidence of which is often associated with considerable morbidity and mortality. Antibiotics that can interact positively with the immune response and that also possess microbicidal properties might significantly contribute to improving the outcome of S. pneumoniae infections. Therefore, in the present study we investigated the effect of clarithromycin, an extended spectrum macrolide currently used in the treatment of respiratory tract infections, on the in vitro interaction between human polymorphonuclear granulocytes (PMN) and three strains of S. pneumoniae with different susceptibility or resistance patterns to both penicillin and clarithromycin. At a concentration of one-half the minimal inhibitory concentration (MIC), clarithromycin significantly enhanced human PMN functions, particularly intracellular bactericidal activity, against all the S. pneumoniae strains, including resistant ones. This finding may help to explain clarithromycin activity in vivo despite apparent resistance in vitro. Preexposure of PMNs to one-half the MIC of clarithromycin had no effect on either phagocytosis or intracellular killing, ruling out a direct antibiotic action on PMNs. Preexposure of streptococci to clarithromycin increased the susceptibility of S. pneumoniae to the bactericidal mechanisms of human PMNs compared with untreated bacteria, indicating that this macrolide may partly reduce bacterial virulence via changes in S. pneumoniae.