Comparative Fungicidal Activities of N-Chlorotaurine and Conventional Antiseptics against Candida spp. Isolated from Vulvovaginal Candidiasis.

N-chlorotaurine (NCT), the N-chloro derivative of the amino acid taurine, is a long-lived oxidant produced by stimulated human leucocytes. NCT has antimicrobial activities which are generally enhanced in the presence of organic material. The aim of this study was to investigate fungicidal effects of NCT and conventional antiseptics against Candida isolated from vulvovaginal candidiasis (VVC). Chlorhexidine (CHX, 1.6%), octenidine dihydrochloride (OCT, 0.08%), povidone iodine (PVP-I, 8%), boric acid (8%), and NCT (0.1% (5.5 mM)) were evaluated against forty-four Candida isolates, according to European Standard methods, at 30, 60, 90, and 120 min and 24 h in the presence of skim milk as an organic material. CHX, OCT, and PVP-I showed rapid fungicidal activity against all Candida isolates with 5-6 log10 reduction of viable counts after 30 min, whereas boric acid and NCT needed 1 h against Candida albicans and 2 h against non-albicans Candida for a significant 3 log10 reduction. NCT showed fungicidal activity (defined as ≥4 log10 reduction) against C. albicans within 90 min and C. non-albicans within 24 h. Based upon all presently available data, including our results, NCT could be used as a new agent for treatment of local fungal infections such as VVC.

Current Opinion on the Therapeutic Capacity of Taurine-Containing Halogen Derivatives in Infectious and Inflammatory Diseases.

Taurine haloamines, N-chlorotaurine (NCT, TauCl), and N-bromotaurine (NBT, TauBr) are formed by a reaction between taurine and hypohalous acids, HOCl and HOBr, respectively. The major source of endogenous taurine haloamines is neutrophils. Both NCT and NBT share strong anti-inflammatory and microbicidal activities supported by an absence of microbial resistance. In the light of these properties, a number of clinical studies have been performed to document their effectiveness in treatment of bacterial, fungal, and viral infections. The administration of NCT and NBT has been limited to topical application, as they are decomposed upon systemic delivery. This review summarizes current knowledge concerning the therapeutic use of NCT and NBT mainly in various skin disorders such as non-healing wounds, acne vulgaris, herpes zoster, and psoriasis. Moreover, the beneficial effect of NCT inhalation in early stages of COVID-19 and other viral respiratory infections is discussed. And finally, we would like to suggest that NCT might be used to inhibit the development of the cytokine storm through its capacity to suppress the production of IL-6.

Taurine and N-Bromotaurine in Topical Treatment of Psoriasis.

Psoriasis is a chronic skin auto-inflammatory and systemic disorder. Novel treatments are needed to solve a plethora of cases refractory to current treatment regimens. N-bromotaurine (TauNH-Br), a natural taurine oxidizing derivative produced by inflammatory cells, has anti-inflammatory, antiproliferative, and antimicrobial properties. This evidence prompted us to use TauNH-Br as a local agent for treatment of therapy-refractory psoriasis. Two pustular-plaque psoriasis cases, unresponsive to systemic and local treatments, one with localized lesions and one with generalized lesions, were selected. Both applications primarily indicated a sufficient curative activity of 1% TauNH-Br in psoriasis lesions. Moreover, TauNH-Br co-administration with taurine and a novel olive oil formulation cut in half the time needed for TauNH-Br alone to cause the same regression of equivalent psoriasis plaque lesions in the same patient. Importantly, all adverse effects of TauNH-Br (erythema, itching, bleeding) could be minimized by the combination therapy. Periods of 2-7 weeks to achieve almost complete regression with this formulation were remarkably short as compared to conventional treatment regimens that both patients had followed previously. Of note, there was no relapse within 3 months of monitoring. Combination formulations containing TauNH-Br and olive oil could become an advantageous topical medication for treatment of psoriasis.

Efficacy of Inhaled N-Chlorotaurine in a Mouse Model of Lichtheimia corymbifera and Aspergillus fumigatus Pneumonia.

N-chlorotaurine (NCT) can be used topically as a well-tolerated anti-infective at different body sites. The aim of this study was to investigate the efficacy of inhaled NCT in a mouse model of fungal pneumonia. Specific pathogen-free female C57BL/6JRj seven-week-old mice were immune-suppressed with cyclophosphamide. After 4 days, the mice were inoculated intranasally with 1.5 × 10E7 spores of Lichtheimia corymbifera or 1.0 × 10E7 spores of Aspergillus fumigatus. They were randomized and treated three times daily for 10 min with aerosolized 1% NCT or 0.9% sodium chloride starting 1 h after the inoculation. The mice were observed for survival for two weeks, and fungal load, blood inflammation parameters, bronchoalveolar lavage, and histology of organs were evaluated upon their death or at the end of this period. Inhalations were well-tolerated. After challenge with L. corymbifera, seven out of the nine mice (77.8%) survived for 15 days in the test group, which was in strong contrast to one out of the nine mice (11.1%) in the control group (p = 0.0049). The count of colony-forming units in the homogenized lung tissues came to 1.60 (1.30; 1.99; median, quartiles) log10 in the test group and to 4.26 (2.17; 4.53) log10 in the control group (p = 0.0032). Body weight and temperature, white blood count, and haptoglobin significantly improved with NCT treatment. With A. fumigatus, all the mice except for one in the test group died within 4 days without a significant difference from the control group. Inhaled NCT applied early demonstrated a highly significant curative effect in L. corymbifera pneumonia, while this could not be shown in A. fumigatus pneumonia, probably due to a too high inoculum. Nevertheless, this study for the first time disclosed efficacy of NCT in pneumonia in vivo.

N-chlorotaurine is highly active against respiratory viruses including SARS-CoV-2 (COVID-19) in vitro.

N-chlorotaurine (NCT) a long-lived oxidant generated by leukocytes, can be synthesized chemically and applied topically as an anti-infective to different body sites, including the lung via inhalation. Here, we demonstrate the activity of NCT against viruses causing acute respiratory tract infections, namely severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza viruses, and respiratory syncytial virus (RSV). Virucidal activity of NCT was tested in plaque assays, confirmed by RT-qPCR assays. Attack on virus proteins was investigated by mass spectrometry. NCT revealed broad virucidal activity against all viruses tested at 37°C and pH 7. A significant reduction in infectious particles of SARS-CoV-2 isolates from early 2020 by 1 log10 was detected after 15 min of incubation in 1% NCT. Proteinaceous material simulating body fluids enhanced this activity by transchlorination mechanisms (1 -2 log10 reduction within 1-10 min). Tested SARS-CoV-2 variants B.1.1.7 (Alpha) und B.1.351 (Beta) showed a similar susceptibility. Influenza virus infectious particles were reduced by 3 log10 (H3N2) to 5 log10 (H1N1pdm), RSV by 4 log10 within a few min. Mass spectrometry of NCT-treated SARS-CoV-2 spike protein and 3C-like protease, influenza virus haemagglutinin and neuraminidase, and RSV fusion glycoprotein disclosed multiple sites of chlorination and oxidation as the molecular mechanism of action. Application of 1.0% NCT as a prophylactic and therapeutic strategy against acute viral respiratory tract infections deserves comprehensive clinical investigation.

Aspergillus terreus and the Interplay with Amphotericin B: from Resistance to Tolerance?

Aspergillus terreus is an opportunistic causative agent of invasive aspergillosis and, in most cases, it is refractory to amphotericin B (AMB) therapy. Notably, AMB-susceptible Aspergillus terreus sensu stricto (s.s.) representatives exist which are also associated with poor clinical outcomes. Such findings may be attributable to drug tolerance, which is not detectable by antifungal susceptibility testing. Here, we tested in vitro antifungal susceptibility (AFST) and the fungicidal activity of AMB against 100 clinical isolates of A. terreus species complex in RPMI 1640 and antibiotic medium 3 (AM3). MICs ranged from 0.5 to 16 µg/mL for RPMI 1640 and from 1 to >16 mg/L for AM3. AMB showed medium-dependent activity, with fungicidal effects only in antibiotic medium 3, not in RPMI 1640. Furthermore, the presence of AMB-tolerant phenotypes of A. terreus has been examined by assessing the minimum duration for killing 99% of the population (MDK99) and evaluating the data obtained in a Galleria mellonella infection model. A time-kill curve analysis revealed that A. terreus with AMB MICs of ≤1 mg/L (susceptible range) displayed AMB-tolerant phenotypes, exhibiting MDK99s at 18 and 36 h, respectively. Survival rates of infected G. mellonella highlighted that AMB was effective against susceptible A. terreus isolates, but not against tolerant or resistant isolates. Our analysis reveals that A. terreus isolates which are defined as susceptible based on MIC may comprise tolerant phenotypes, which may, in turn, explain the worse outcome of AMB therapy for phenotypically susceptible isolates.

Recombinant actin-depolymerizing factor of the apicomplexan Neospora caninum (NcADF) is susceptible to oxidation.

Neospora caninum is a member of Apicomplexa Phylum and the causative agent of neosporosis, a disease responsible for abortions in cattle. Apicomplexan parasites have a limited set of actin-binding proteins conducting the regulation of the dynamics of nonconventional actin. The parasite actin-based motility is implicated in the parasite invasion process in the host cell. Once no commercial strategy for the neosporosis control is available, the interference in the parasite actin function may result in novel drug targets. Actin-depolymerization factor (ADF) is a member of the ADF/cofilin family, primarily known for its function in actin severing and depolymerization. ADF/cofilins are versatile proteins modulated by different mechanisms, including reduction and oxidation. In apicomplexan parasites, the mechanisms involved in the modulation of ADF function are barely explored and the effects of oxidation in the protein are unknown so far. In this study, we used the oxidants N-chlorotaurine (NCT) and H2O2 to investigate the susceptibility of the recombinant N. caninum ADF (NcADF) to oxidation. After exposing the protein to either NCT or H2O2, the dimerization status and cysteine residue oxidation were determined. Also, the interference of NcADF oxidation in the interaction with actin was assessed. The treatment of the recombinant protein with oxidants reversibly induced the production of dimers, indicating that disulfide bonds between NcADF cysteine residues were formed. In addition, the exposure of NcADF to NCT resulted in more efficient oxidation of the cysteine residues compared to H2O2. Finally, the oxidation of NcADF by NCT reduced the ability of actin-binding and altered the function of NcADF in actin polymerization. Altogether, our results clearly show that recombinant NcADF is sensitive to redox conditions, indicating that the function of this protein in cellular processes involving actin dynamics may be modulated by oxidation.

Activity of N-Chlorotaurine against Long-Term Biofilms of Bacteria and Yeasts.

Background: N-chlorotaurine (NCT), an antiseptic that originates from the human defense system, has broad-spectrum microbicidal activity and is well tolerated by human tissue and applicable to sensitive body regions. Bacteria in short-term biofilms, too, have been shown to be killed by NCT. It was the aim of the present study to demonstrate the activity of NCT against bacteria and yeasts in longer-lasting biofilms, including their co-culture. Materials and methods: Staphylococcus aureus, Pseudomonas aeruginosa, and Klebsiella variicola biofilms were grown for 14 weeks in MBECTM inoculator with 96 well base. Some pegs were pinched off weekly and incubated in 1% NCT in PBS (PBS only for controls) at pH 7.1 and 37 °C, for 30 and 60 min. Subsequently, bacteria were resuspended by ultrasonication and subjected to quantitative cultures. Similar tests were conducted with C. albicans biofilms grown on metal (A2-steel) discs for 4 weeks. Mixed co-cultures of C. albicans plus each of the three bacterial strains on metal discs were grown for 5-7 weeks and weekly evaluated, as mentioned above. Results: Single biofilms of S. aureus, P. aeruginosa, and K. variicola grew to approximately 1 × 106 colony forming units (CFU)/mL and C. albicans to 1 × 105 CFU/mL. In combined biofilms, the CFU count was about 1 log10 lower. Viable counts of biofilms of single bacteria were reduced by 2.8 to 5.6 log10 in 1% NCT after 60 min (0.9 to 4.7 log10 after 30 min) with Gram-negative bacteria being more susceptible than S. aureus. Significant reduction of C. albicans by 2.0 to 2.9 log10 occurred after 4 h incubation. In combined biofilms, viable counts of C. albicans were reduced by 1.1 to 2.4 log10 after 4 h, while they reached the detection limit after 1 to 2 h with bacteria (2.0 to > 3.5 log10 reduction). Remarkably, older biofilms demonstrated no increase in resistance but constant susceptibility to NCT. This was valid for all tested pathogens. In electron microscopy, morphological differences between NCT-treated and non-treated biofilms could be found. Conclusions: NCT is active against long-term biofilms of up to several months irrespective of their age. Combined biofilm cultures of yeasts and bacteria show a similar susceptibility pattern to NCT as single ones. These results contribute to the explanation of the clinical efficacy of NCT, for instance, in infected chronic wounds and purulently coated crural ulcerations.

Oxidant therapy improves adipogenic differentiation of adipose-derived stem cells in human wound healing.

BACKGROUND: Adipose-derived stem cells (ASC) and adipocytes are involved in numerous physiological and pathophysiological conditions, which have been extensively described in subcutaneous and visceral fat depots over the past two decades. However, much less is known about ASC and adipocytes outside classical fat tissue depots and their necessity in tissue remodeling after injury. Therefore, we investigated the etiology of adipocytes in human granulation tissue and define their possible role wound healing. METHODS: Identification of human wound tissue adipocytes was determined by immunohistochemical staining of granulation tissue sections from patients undergoing surgical debridement. Stromal cell fractions from granulation tissue and subcutaneous fat tissue were generated by collagenase type II-based protocols. Pro- and anti-inflammatory wound bed conditions were mimicked by THP1- and CD14+ monocyte-derived macrophage models in vitro. Effects of macrophage secretome on ASC differentiation and metabolism were determined by immunoblotting, flow cytometry, and microscopy assessing early and late adipocyte differentiation states. Functional rescuing experiments were conducted by lentiviral transduction of wildtype PPARG, IL1RA, and N-chlorotaurine (NCT) treatment. RESULTS: Single and clustered adipocyte populations were detected in 11 out of 13 granulation tissue specimens and single-cell suspensions from granulation tissue showed adipogenic differentiation potential. Pro-inflammatory signaling by IFNG/LPS-stimulated macrophages (M (IFNG/LPS)) inhibited the maturation of lipid droplets in differentiated ASC. In contrast, anti-inflammatory IL4/IL13-activated macrophages (M (IL4/IL13)) revealed minor effects on adipocyte development. The M (IFNG/LPS)-induced phenotype was associated with a switch from endogenous fatty acid synthesis to glycolysis-dominated cell metabolism and increased pro-inflammatory cytokine production. Impaired adipogenesis was associated with increased, but seemingly non-functional, CEBPB levels, which failed to induce downstream PPARG and CEBPA. Neither transgenic PPARG overexpression, nor inhibition of IL1B was sufficient to rescue the anti-adipogenic effects induced by IFNG/LPS-activated macrophages. Instead, macrophage co-treatment during stimulation with NCT, a mild oxidant produced by activated granulocytes present in human wounds in vivo, significantly attenuated the anti-adipogenic effects. CONCLUSIONS: In conclusion, the appearance of adipocytes in wound tissue indicates a prevailing anti-inflammatory environment that could be promoted by NCT treatment and may be associated with improved healing outcomes.

Bromamine T (BAT) Exerts Stronger Anti-Cancer Properties than Taurine (Tau).

BACKGROUND: Taurine (Tau) ameliorates cancer pathogenesis. Researchers have focused on the functional properties of bromamine T (BAT), a stable active bromine molecule. Both N-bromotaurine (TauNHBr) and BAT exert potent anti-inflammatory properties, but the landscape remains obscure concerning the anti-cancer effect of BAT. METHODS: We used Crystal Violet, colony formation, flow cytometry and Western blot experiments to evaluate the effect of BAT and Tau on the apoptosis and autophagy of cancer cells. Xenograft experiments were used to determine the in vivo cytotoxicity of either agent. RESULTS: We demonstrated that both BAT and Tau inhibited the growth of human colon, breast, cervical and skin cancer cell lines. Among them, BAT exerted the greatest cytotoxic effect on both RKO and MDA-MB-468 cells. In particular, BAT increased the phosphorylation of c-Jun N-terminal kinases (JNK½), p38 mitogen-activated protein kinase (MAPK), and extracellular-signal-regulated kinases (ERK½), thereby inducing mitochondrial apoptosis and autophagy in RKO cells. In contrast, Tau exerted its cytotoxic effect by upregulating JNK½ forms, thus triggering mitochondrial apoptosis in RKO cells. Accordingly, colon cancer growth was impaired in vivo. CONCLUSIONS: BAT and Tau exerted their anti-tumor properties through the induction of (i) mitochondrial apoptosis, (ii) the MAPK family, and iii) autophagy, providing novel anti-cancer therapeutic modalities.

Successful treatment of a unique chronic multi-bacterial scalp infection with N-chlorotaurine, N-bromotaurine and bromamine T.

Microbial species can act in synergy to circumvent environmental stress conditions and survive. In addition, biofilms are a serious public-health issue globally and constitute a clinical emergency. Infection persistence, increased morbidity and mortality, and antibiotic resistance are consequences of poly-microbial synergy. Due to inherited complexity and synergy between numerous species, newer antimicrobial agents of increased efficacy and tolerability are needed. In this unique medical case, a chronic (9 year) multi-bacterial scalp infection was differentially diagnosed from other inflammatory skin disorders by prolonged microbiological culture. The bacterial species found seem to have caused lesions of visible biofilm not documented previously in the medical literature. This complicated infection was treated successfully and rapidly with the combined topical application of the active halogen compounds N-chlorotaurine, N-bromotaurine and bromamine T, which is in contrast to the previous failed systemic and topical therapeutic approaches. This study strengthens the case for the use of active halogen compounds against multi-bacterial infections of the skin in the future, without the occurrence of resistance.

Microbicidal activity of N-chlorotaurine can be enhanced in the presence of lung epithelial cells.

BACKGROUND: N-chlorotaurine (NCT) is an endogenous active chlorine compound that can be used as an antiseptic and anti-infective in different body regions. Recently, tolerability of inhaled NCT has been demonstrated in humans so that it is of interest for future treatment of cystic fibrosis. In the present study, we tested the bactericidal and fungicidal activity of NCT in different lung cell culture models. METHODS: Bacteria (Staphylococcus aureus, Pseudomonas aeruginosa) and fungi (Candida albicans, Exophiala dermatitidis) were co-incubated with lung epithelial cell cultures, and after 4 h NCT was added. After different incubation times, aliquots were removed and quantitative cultures were performed. RESULTS: NCT at the therapeutically applied concentration of 1% (55 mM) completely killed the test pathogens within 15 - 30 min at 20 °C and at 37 °C. Killing by 0.3% NCT lasted up to 4 h dependent on the pathogen at 20 °C and up to 1 h at 37 °C. 0.1% NCT was the threshold concentration for killing since this amount of oxidation capacity was consumed by reactions with the organic compounds of the medium within 3 h (20 °C) and 0.5 h (37 °C). CONCLUSIONS: NCT in therapeutic concentration demonstrated its microbicidal activity in the presence of lung epithelial cells. Remarkably, particularly the fungicidal activity was higher under these conditions than in phosphate buffer. This can be explained by formation of the stronger microbicidal monochloramine in equilibrium by transchlorination. The results suggest the suitability of NCT as inhalation medication in the lung.

Development and Validation of an HPLC Method for the Quantitative Analysis of Bromophenolic Compounds in the Red Alga Vertebrata lanosa.

Bromophenols are a class of compounds occurring in red algae that are thought to play a role in chemical protection; however, their exact function is still not fully known. In order to investigate their occurrence, pure standards of seven bromophenols were isolated from a methanolic extract of the epiphytic red alga Vertebrata lanosa collected in Brittany, France. The structures of all compounds were determined by NMR and MS. Among the isolated substances, one new natural product, namely, 2-amino-5-(3-(2,3-dibromo-4,5-dihydroxybenzyl)ureido)pentanoic acid was identified. An HPLC method for the separation of all isolated substances was developed using a Phenomenex C8(2) Luna column and a mobile phase comprising 0.05% trifluoroacetic acid in water and acetonitrile. Method validation showed that the applied procedure is selective, linear (R2 0.999), precise (intra-day ≤ 6.28%, inter-day ≤ 5.21%), and accurate (with maximum displacement values of 4.93% for the high spikes, 4.80% for the medium spikes, and 4.30% for the low spikes). For all standards limits of detection (LOD) were lower than 0.04 µg/mL and limits of quantification (LOQ) lower than 0.12 µg/mL. Subsequently, the method was applied to determine the bromophenol content in Vertebrata lanosa samples from varying sampling sites and collection years showing values between 0.678 and 0.005 mg/g dry weight for different bromophenols with significant variations between the sampling years. Bioactivity of seven isolated bromophenols was tested in agar diffusion tests against Staphylococcus aureus and Escherichia coli bacteria. Three compounds showed a small zone of inhibition against both test organisms at a concentration of 100 µg/mL.

Combined Biological Effects of N-Bromotaurine Analogs and Ibuprofen. Part I: Influence on Inflammatory Properties of Macrophages.

Taurine haloamines (N-chlorotaurine, N-bromotaurine) due to their strong antiseptic and anti-inflammatory properties are good candidates for topical application in treatment of skin inflammatory/infectious disorders. Recently, we have demonstrated that more stable N-bromotaurine analogs (N-dibromo-dimethyl taurine, N-monobromo-dimethyl taurine) and bromamine T show strong microbicidal and anti-inflammatory properties at concentrations well tolerated by human cells and tissue. Non-steroidal anti-inflammatory drugs (NSAIDs) with cyclooxygenase (COX) inhibitory activity are commonly used in various inflammatory diseases. However, systemic administration of NSAIDs may result in adverse side effects. For example, the use of ibuprofen in children with varicella is associated with enhanced serum levels of TNF-α and with increased risk of necrotizing soft tissue infections and secondary skin infections caused by invasive streptococci. The aim of this study was to examine combined immunomodulatory effects of bromamines and ibuprofen on J774.A1 macrophages. We have shown that the primary activity of ibuprofen, the inhibition of PGE2 production by activated macrophages was intensified in the presence of bromamines. Most importantly, the stimulatory effect of ibuprofen on production of inflammatory cytokines (TNF-α, IL-6) was inhibited by all tested bromamines. These observations indicate that bromamines may neutralize massive production of TNF-α at sites of inflammation, a side effect of ibuprofen. Therefore, we suggest that systemic administration of ibuprofen (NSAIDs) in treatment of inflammatory/infectious skin diseases should be supported by topical application of bromamines as an adjunctive therapy.

Combined Biological Effects of N-Bromotaurine Analogs and Ibuprofen. Part II: Influence on a Local Defense System.

The stable N-bromotaurine analogs (N-dibromo-dimethyl taurine, N-monobromo-dimethyl taurine), and bromamine T (BAT) show anti-inflammatory and microbicidal properties. These bromamines are good candidates for a treatment of skin infectious/inflammatory diseases as local antiseptics. Ibuprofen, a non-steroidal anti-inflammatory drug (NSAID), is commonly used in various infectious/inflammatory diseases due to its analgesic and antipyretic therapeutic effects. However, systemic administration of ibuprofen may also result in adverse side effects. It has been reported that ibuprofen enhances serum levels of TNF-α and worsens secondary skin infections caused by invasive streptococci (S. pyogenes). Recently we have demonstrated that bromamines inhibit the stimulatory effect of ibuprofen on the production of inflammatory cytokines (TNF-α, IL-6). The aim of this study was to examine the combined antibacterial actions of ibuprofen and bromamines against S. pyogenes and their joint effect on the generation of reactive oxygen species (ROS) by activated neutrophils and macrophages. We have shown that the microbicidal activity of bromamines against S. pyogenes was not altered by ibuprofen. On the other hand, co-administration of ibuprofen and bromamines markedly decreased the generation of ROS by activated neutrophils and macrophages. Finally, we discuss how the antioxidant combined effect of bromamines and ibuprofen may affect a local defense system.

N-chlorotaurine Inactivates Acanthamoeba and Candida albicans in the Porcine Ex Vivo Corneal Infection Model.

PURPOSE: N-chlorotaurine (NCT) is an anti-infective belonging to the class of chloramines and an investigative drug for the topical treatment of keratoconjunctivitis. The aim of the present study was to demonstrate its efficacy against Acanthamoeba and Candida in corneas infected ex vivo. METHODS: Corneal buttons from porcine eyes were contaminated with Acanthamoeba castellanii trophozoites or Candida albicans Centraalbureau voor Schimmelcultures 5982 and incubated for 7 and 3 days, respectively. Subsequently, they were treated with 1% NCT for 5 to 120 minutes. After further incubation for 2 days in the absence of NCT in tests with A. castellanii, the buttons were homogenized, and the amoebae grown for a further 5 days before they were counted in a light microscope. For C. albicans, quantitative cultures were performed from corneal homogenates. RESULTS: Incubation of 120 minutes in NCT completely inhibited the regrowth of A. castellanii and reduced the number of C. albicans colony-forming unit counts by 4 log10. In addition, at 60 minutes, significant reductions of both pathogens could be observed. Histology showed penetration of pathogens into the stroma of the corneal buttons. CONCLUSIONS: NCT inactivates A. castellanii and C. albicans in corneal tissue.

Swift Cure of a Chronic Wound Infected With Multiresistant Staphylococcus aureus in an Elderly Patient With Stage 5 Renal Disease.

We present a case of a 91-year-old female with stage 5 renal disease, diabetes type 2, and considerable weakness, suffering from a 2-month-old wound infected by a multiresistant Staphylococcus aureus. The wound measured 7 cm in length, 5 cm in width, and 1.5 cm in depth, having purulent white edges and exudates exceeding the size of the wound. The systemic antibiotic use was opposing to improve the patient's clinical condition due to underlying nephrotoxicity that may have deteriorated renal failure and resistance of the infecting pathogen. The halogenated taurine (Tau) derivatives N-chlorotaurine (NCT) and N-bromotaurine (NBrT) with potent anti-inflammatory and antimicrobial efficacy were alternatively employed as combination topical treatment to provide a therapeutic solution. Each agent was applied separately with an interval of 5 minutes as a 1% spray in aqueous solution every 30 minutes during the day for 3 days. This treatment was very well tolerated and led to rapid disappearance of the purulent exudate, rapid epithelialization, and complete healing. To avoid relapse, the application was continued 4 times daily for a further 4 days. No complications occurred in the course of treatment. This case report confirms the therapeutic efficacy of NCT in chronic purulent wounds. NBrT is well tolerated, too, and can be used in combination with NCT in emergency clinical settings. Its potential as a single agent should be investigated in further studies. Advancement of wound closure by these agents proved to be life-saving for this patient. Further molecular research is needed to identify mechanisms that promote wound healing.

Inactivation of Adenovirus in Water by Natural and Synthetic Compounds.

Millions of people use contaminated water sources for direct consumption. Chlorine is the most widely disinfection product but can produce toxic by-products. In this context, natural and synthetic compounds can be an alternative to water disinfection. Therefore, the aim of this study was to assess the inactivation of human adenovirus by N-chlorotaurine (NCT), bromamine-T (BAT) and Grape seed extract (GSE) in water. Distilled water artificially contaminated with recombinant human adenovirus type 5 (rAdV-GFP) was treated with different concentrations of each compound for up to 120 min, and viral infectivity was assessed by fluorescence microscopy. The decrease in activity of the compounds in the presence of organic matter was evaluated in water supplemented with peptone. As results, NCT and GSE inactivated approximately 2.5 log10 of adenovirus after 120 min. With BAT, more than 4.0 log10 decrease was observed within 10 min. The oxidative activity of 1% BAT decreased by 50% in 0.5% peptone within a few minutes, while the reduction was only 30% for 1% NCT in 5% peptone after 60 min. Organic matter had no effect on the activity of GSE. Moreover, the minimal concentration of BAT and GSE to kill viruses was lower than that known to kill human cells. It was concluded that the three compounds have potential to be used for water disinfection for drinking or reuse purposes.

Exposure to N-chlorotaurine induces oxidative stress responses in Aspergillus fumigatus.

PURPOSE: The neutrophil-derived oxidant N-chlorotaurine (NCT) displays remarkable in vivo tolerability and efficacy against a range of pathogens. The aim of this study was to characterize the response of the pulmonary pathogen Aspergillus fumigatus to NCT. METHODOLOGY: The effect of NCT on the growth and viability of A. fumigatus was characterized. NCT-induced alteration of amino acids and gliotoxin from A. fumigatus mycelium was assessed. Label-free shotgun quantitative proteomic analysis was performed on A. fumigatus exposed to NCT for 24 h. RESULTS: Incubation of A. fumigatus with NCT at concentrations ranging from 6.8 to 55 mM decreased conidial growth and viability, and mycelium biomass relative to the controls. Exposure to NCT (13.77 mM) resulted in increased amino acids and gliotoxin levels from A. fumigatus mycelium. Exposure of A. fumigatus mycelium to NCT (6.8 mM) revealed an enrichment in proteins associated with the ribosome, transcription and translation and non-ribosomal peptide biosynthesis (e.g. Pes1, Pes3), which play an essential role in oxidative stress resistance in A. fumigatus. A decrease in the abundance of proteins associated with fumagillin and pseurotin biosynthesis highlighted the anti-virulence activity of NCT. CONCLUSION: These results indicate that NCT induces an oxidative stress response in A. fumigatus as evidenced by alterations in the proteome and inhibits conidial and mycelial growth. Clinical investigations of topical application of NCT to treat Aspergillus infections are encouraged.

Potentiometric characterization of iodine-based disinfecting preparations.

A potentiometric method is presented that allows to characterize aqueous iodine-based disinfecting solutions and preparations, respectively. By means of three electrodes (iodide-sensitive, platinum redox, and reference electrode), the concentrations of free iodine, iodide, and triiodide were determined. The apposition "free" means iodine not complexed with ligands originating from the organic matrix of the iodophoric iodine source (e.g. povidone iodine). Based on the gained values ([I2], [I-], and[I3-]), it is possible to assess features concerning kinetics of disinfection, disinfecting capacity, and tolerability. The method distinguishes by simplicity and modest time expenditure of 10-15 min for a complete investigation comprising two potentiometric measurements.