The small molecule, thiazolidinedione-8 (S-8) was shown to impair biofilm formation of various microbial pathogens, including the fungus Candida albicans and Streptococcus mutans. Previously, we have evaluated the specific molecular mode of S-8 action against C. albicans biofilm-associated pathogenicity. In this study we investigated the influence of S-8 on dual species, C. albicans-S. mutans biofilm. We show that in the presence of S-8 a reduction of the co-species biofilm formation occurred with a major effect on C. albicans. Biofilm biomass and exopolysaccharide (EPS) production were significantly reduced by S-8. Moreover, the agent caused oxidative stress associated with a strong induction of reactive oxygen species and hydrogen peroxide uptake inhibition by a mixed biofilm. In addition, S-8 altered symbiotic relationship between these species by a complex mechanism. Streptococcal genes associated with quorum sensing (QS) (comDE and luxS), EPS production (gtfBCD and gbpB), as well as genes related to protection against oxidative stress (nox and sodA) were markedly upregulated by S-8. In contrast, fungal genes related to hyphae formation (hwp1), adhesion (als3), hydrophobicity (csh1), and oxidative stress response (sod1, sod2, and cat1) were downregulated in the presence of S-8. In addition, ywp1 gene associated with yeast form of C. albicans was induced by S-8, which is correlated with appearance of mostly yeast cells in S-8 treated dual species biofilms. We concluded that S-8 disturbs symbiotic balance between C. albicans and S. mutans in dual species biofilm.

In 2009, Xu et al. and Chaput et al. in Nature Medicine had argued that the main cause of death in sepsis is the release from neutrophil nets of nuclear histone, highly toxic to endothelial cells and that these polycations are major and unique virulence factors. Since 2009, numerous researchers have also suggested the involvement of histones in the pathophysiology of many clinical disorders. If histones are indeed major unique virulence toxic agents, then heparin, activated protein C and antibodies to histone should prove excellent antisepsis agents. However, this is provided that these agents are administered to patients early enough before the activation of the cytokine storms, immune responses and the coagulation cascades are irreversibly unleashed. This may not be practical, since a diagnosis of sepsis is usually made much later. Future identifications of novel early markers are therefore needed and a compilation of cocktails of antagonists may replace the faulty single antagonists tried for many years, but in vain, to prevent death in sepsis.

It is disconcerting and also alarming that today clinicians are still bewildered and helpless when trying to cope with life-threatening sequelae of severe microbial infections, which very often culminate in sepsis, septic shock and death. According to CDC (the Centers for Disease Control and Prevention), the annual incidence of sepsis in the USA affects as many as 750,000 hospitalized patients and mortality rate is about 40% [1,2]. It was found in 2 complementary inpatient cohorts that up to 50% of hospital deaths were linked to sepsis [3]. Worldwide, sepsis is one of the common deadly diseases. It is one of the few conditions to strike with equal ferocity in resource-poor areas and in the developed world. Globally, 20 to 30 million patients are estimated to be afflicted every year. Every hour, about 1,000 people and each day around 24,000 people die from sepsis worldwide and sepsis is one of the least well known diseases. In the developing world, sepsis accounts for 60-80% of lost lives in childhood, with more than 6 million neonates and children affected by sepsis annually. Sepsis is responsible for >100,000 cases of maternal sepsis each year and in some countries is now a greater threat in pregnancy than bleeding or thromboembolism [4,5]. Screening the voluminous literature on sepsis treatment revealed unsuccessful efforts to save patients' lives by administering antibiotics but only a signally-chosen antagonist at a time. The numbers of anti-inflammatory agents tested ineffectively over the years is phenomenal (see below) and today even the most promising activated protein C, the “miracle drug” was recently discontinued [6-9]. The initial reactions to infection are generalized pro-and anti-inflammatory responses. These usually starts by activation by microorganisms and some of their products of neutrophils, macrophages and monocytes, which are followed by toxic effects on vascular endothelial cells via pathogen recognition receptors, leading to endothelial disruption. Why have all the therapeutic strategies tested invariably failed to cope with the sequelae of severe microbial infections and what future approaches might break the stalemate leading to a better understanding of the pathophysiology of the "horror autotoxicus" phenomena of sepsis? [10].

Reviewing the “glorious history” of medical microbiology revealed that immunoglobulins rich in anti-toxins activities proved very effective to cope with those maladies where a single virulence agent, such as the toxin of diphtheria, tetanus and botulism, are the main pathogenetic virulence agents. Also, anti-viral vaccines are the hallmark of the prevention of many childes viral diseases and of viral hepatitis. On the other hand, no single major virulence factor is identified in the majority of Gram positives Gram negatives, fungal and Mycobacterial pathogens. Therefore, it stands to reason that cell and tissue damage inflicted by these microorganisms may be a result of a coordinated "cross-talk" (synergism) among host factors and a multiplicity of pro-inflammatory agents generated during the proliferation of bacteria, mainly in the blood stream. These may include: extracellular pore-forming and membrane-permeabilizing hemolysins, capsular polysaccharides, LPS (endotoxin), the membrane-associated lipoteichoic acid (LTA), the rigid cell-wall peptidoglycan (PPG), leukocyte-derived oxygen and nitrogen species, anti-microbial cationic peptides, phospholipases, cationic proteinases, growth factors, cytokines and chemokines and many others. All these agents might be generated in various stages of inflammation and infection by microbes and by the host response. Furthermore, certain life-saving antibiotics might also act as "double-edged swords" by enhancing the release of microbial products (LPS, LTA, PPG, capsular polysaccharides, intra cellular toxins), resulting from to the activation of nascent autolytic wall enzymes released leading to bacteriolysis [11,12].

The oxidant scavenging ability (OSA) of catalase-rich Candida albicans is markedly enhanced by chlorhexidine digluconate (CHX), polymyxin B, the bile salt ursodeoxycholate and by lysophosphatidylcholine, which all act as detergents facilitating the penetration of oxidants and their intracellular decomposition. Quantifications of the OSA of Candida albicans were measured by a highly sensitive luminol-dependent chemiluminescence assay and by the Thurman's assay, to quantify hydrogen peroxide (H2O2). The OSA enhancing activity by CHX depends to some extent on the media on which candida grew. The OSA of candida treated by CHX was modulated by whole human saliva, red blood cells, lysozyme, cationic peptides and by polyphenols. Concentrations of CHX, which killed over 95 % of Candida albicans cells, did not affect the cells' abilities to scavenge reactive oxygen species (ROS). The OSA of Candida cells treated by CHX is highly refractory to H2O2 (50 mM) but is strongly inhibited by hypochlorous acid, lecithin, trypan blue and by heparin. We speculate that similarly to catalase-rich red blood cells, Candida albicans and additional catalase-rich microbiota may also have the ability to scavenge oxidants and thus can protect catalase-negative anaerobes and facultative anaerobes cariogenic streptococci against peroxide and thus secure their survival in the oral cavity.

tives and their involvement in the pathogenesis of chronic granulomatous inflammation is briefly reviewed. It can be speculated that in humans, leukocytes laden with intracellular bacteria and their non-degraded highly-phlogistic cell-walls may be translocated from inflamed gums (periodontal disease) and from infected dental pulps (pulpitis, periapical granulomas) to remote sites such as damaged heart valves (causing endocarditis) and injured joints (causing chronic arthritis). This phenomenon maybe important, clinically and is in line with the old “Focus of infection theory” from the nineteen twenties, which is no longer considered and discussed in the modern literature.

Oxidative stress has been recognized to play important roles in various diseases, including of the oral cavity. However, nutritional supplementation of antioxidants to ameliorate the consequences of oxidative stress is debatable. One caveat is that oxidative status is often measured under non-physiological conditions. Here, we investigated the antioxidant potential of fermented papaya preparation (FPP), a product of yeast fermentation of Carica papaya Linn, under conditions that prevail in the oral cavity. Employing highly sensitive luminol-dependent chemiluminescence assays, we show that its antioxidant capacity was augmented by saliva (up to 20-fold, p<0.0001, at 10mg) and its components (mucin, albumin) as well as by red blood cells (RBC) and microorganisms present in the normal and pathological environment of the oral cavity. Polyphenols are major plant antioxidants. Using the Folin–Ciocalteu’s assay, a very low amount of phenols was measured in FPP suspended in a salt solution. However, its suspension in saliva, albumin, mucin or RBC produced up to sixfold increase, p<0.001, compared with the sum of polyphenols assayed separately. The results suggested that these enhancing effects were due to the solubilization of antioxidant polyphenols in FPP by saliva proteins and the binding to RBC and microorganisms, thus increasing their availability and activity. Copyright © 2015 John Wiley & Sons, Ltd.

LETTER Here are some comments and useful suggestions after reading an article in mBio by Brown et al. entitled “Mechanisms underlying the exquisite sensitivity of Candida albicans to combinatorial cationic and oxidative stress that enhances the potent fungicidal activity of phagocytes” (1). In this paper, we are informed that a simultaneous exposure to 5 mM H2O2 and to cationic NaCl at 1 M is much more potent than the individual stresses themselves and that this combinatorial stress kills C. albicans synergistically in vitro. Such combinations are obviously absolutely unrealistic and not physiological. As a comparison I wonder why the authors had not also tested naturally occurring antimicrobial cationic peptides such as LL37 found in large amounts in neutrophil granules? Had the authors read the classical papers describing the possible mechanisms of bactericidal effects of neutrophils, they would have realized that there is actually no free-floating H2O2 in phagosomes following phagocytosis. This is because activation of NADPH-oxidase yields superoxide, which very rapidly interacts with myeloperoxidase (MPO) and with a halide (Cl−) to generate microbicidal amounts of hypochlorous acid (HOCl) (2–5)! Therefore, HOCl should have definitely been considered and tested in the system described by the authors. Also, the term flux used may be inappropriate since, in their study, both H2O2 and NaCl were actually applied as a bolus. Fluxes of oxidants are generated mainly by activated neutrophils and macrophages and by xanthine and xanthine oxidase in endothelial cells (2) Also, I wonder whether Na used is specific and whether potassium ions can also have the same effects in their system? The authors also claimed that catalase-derived peroxide detoxification, which is inhibited by cations, leads to intracellular ROS accumulation because catalase activity had been affected. If so, why had the catalase inhibitor azide or aminotriazole not been tested? In their study, the authors grew Candida cells in Tris-buffered yeast extract-peptone-dextrose medium (YPDT; pH 7.4). However, the authors have not cited key papers showing that D-glucose, in media on which candida grow, may also suppress catalase formation (6, 7). Using unrealistic, nonphysiological amounts of reagents will not increase our understanding of how biological processes really occur in vivo, despite the need to employ in vitro models. Also, disregarding key published data on neutrophil functions and Candida biology is unacceptable. Can this be a “menace to the future of honest science” (8) and also a “transgression” (9)? See also a recent publication by Casadevall and Fang (10).