Violacein and its antifungal activity: comments and potentialities.

Violacein is an important natural antimicrobial pigment that is mainly produced by Chromobacterium violaceum and Janthinobacterium lividum. It presents a significant range of effects against phytopathogenic and human fungi, besides being featured as having low toxicity, and by its important ecological role in protecting amphibian species and applications in dyed medical fabric. The hypothesis about violacein's action mechanisms against mucormycosis (Rhizopus arrhizus) and candidiasis (Candida auris) is herein discussed based on data available in the scientific literature.

The nanotechnology among US: are metal and metal oxides nanoparticles a nano or mega risk for soil microbial communities?

Metal nanoparticles and metal oxides nanoparticles (MNPs/MONPs) have been widely included in a great diversity of products and industrial applications and they are already a part of our everyday life. According to estimation studies, their production is expected to increase exponentially in the next few years. Consequently, soil has been suggested as the main sink of MNPs/MONPs once they are deliberately or accidentally released into the environment. The potential negative perturbations that may result on soil microbial communities and ecological processes are resulting in concerns. Several nano-toxicological studies of MNPs/MONPs, reported so far, have focused on aquatic organisms, animals, and soil invertebrates. However, during recent years, the studies have been oriented to understand the effects of MNPs/MONPs on microbial communities and their interaction with soil components. The studies have suggested that MNPs/MONPs are one of the most toxic type to soil biota, amongst different types of nanomaterials. This may threaten soil health and fertility, since microbial communities are known to support important biological processes and ecosystem services such as the nutrient cycling, whereby their protection against the environmental pollution is imperative. Therefore, in this review we summarize the actual knowledge available from the last five years (2013-2018) and gaps about the potential negative, positive or neutral effects produced on soil by different classes of MNPs/MONPs. A particular emphasis has been placed on the associated soil microorganisms and biological processes. Finally, perspectives about future research are discussed.

In vitro investigation of antimicrobial effects, nanohardness, and cytotoxicity of different glass ionomer restorative materials in dentistry.

AIMS: The aim of this in vitro study was to investigate the antimicrobial effects, nanohardness, and cytotoxicity of different glass-ionomer restorative materials in dentistry. MATERIALS AND METHODS: The following glass-ionomer restorative materials were used in our study: Argion (VOCO, Germany), Zirconomer (Shofu Inc., Japan), EQUIA Forte (GC, Japan), Fuji II LC capsule (GC, Japan), and Fuji IX GP capsule (GC, Japan). For the cytotoxicity test, a cell culture without release liquids was used as the control group. Microdilution and a disk diffusion test were used to measure the antimicrobial activity. The MTT (3- [4,5-dimethylthiazol-2-yl] -2,5-diphenyltetrazolium bromide) assay was used to evaluate cytotoxicity. Nanohardness was measured using a nanoindenter device. RESULTS: Our study determined that all restorative materials used in this study inhibited bacterial growth in cultures containing 1 × 102 cfu/ml of the bacterial concentrations (Lactobacillus casei and Streptococcus mutans) and that all restorative materials inhibited fungal growth in the cultures containing <1 × 102 cfu/ml of the fungal strains (Candida albicans). IC50 values obtained for the cytotoxicity test were analyzed using the Chi-square test. After a 72-h incubation period, Zirconomer, EQUIA Forte, Fuji IX GP capsule, and Fuji II LG capsule showed statistically significant lower cell viability values. (P < 0.05). The Kruskal-Wallis analysis was performed on the values obtained from the nanohardness test; the differences between the groups were found to be significant (P < 0.05). CONCLUSIONS: All materials tested showed low antimicrobial activity, and the antifungal activity of these materials was found to be lower than their antimicrobial activity. Only Argion showed non-cytotoxic effect. Beginning with the group with the highest nanohardness values and ending with the lowest, the groups are ranked in the following order: Fuji II LC capsule, EQUIA Forte, Argion, Fuji IX GP capsule, and Zirconomer.

Are white-rot fungi a real biotechnological option for the improvement of environmental health?

The use of white-rot fungi as a biotechnological tool for cleaning the environment of recalcitrant pollutants has been under evaluation for several years. However, it is still not possible to find sufficiently detailed investigations of this subject to conclude that these fungi can decontaminate the environment. In the present review, we have summarized and discussed evidence about the potential of white-rot fungi to degrade such pollutants as polycyclic aromatic hydrocarbons, dyes or antibiotics as an example of the complex structures that these microorganisms can attack. This review also discusses field experiment results and limitations of white-rot fungi trials from contaminated sites. Moreover, the use of catabolic potential of white-rot fungi in biopurification systems (biobeds) is also discussed. The current status and future perspectives of white-rot fungi, as a viable biotechnological alternative for improvement of environmental health are noted.

Beta-lactamase Enzymes of Clinical Pseudomonas aeruginosa Strains.

OBJECTIVES: In this study, the production of extended spectrum beta-lactamase (ESBL), metallo-betalacatamase (MBL) and AmpC beta-lactamase enzymes of Pseudomonas aeruginosa (P aeruginosa) strains which were isolated from clinical samples were investigated. AmpC gene was also detected by the polymerase chain reaction (PCR) analysis. METHODS: A hundred strains of P aeruginosa were included in the study. The presence of ESBL was investigated with combined disk confirmation test, MBL was investigated with E-test method and AmpC beta-lactamase was investigated with disk induction test. In order to detect the production of AmpC betalactamase genotypically, the PCR method was used. RESULTS: Only one strain was found to be MBL positive. Four per cent of strains were found to be ESBL positive. AmpC beta-lactamase production was positive in 73% of the strains with disk induction test. AmpC gene was detected in 96% of the studied strains with the PCR method. CONCLUSION: While ESBL and MBL rates in this study were significantly lower than those found in other studies, the rate of AmpC beta-lactamase was higher. Although AmpC gene was detected in some strains (23%), they were not found to produce AmpC beta-lactamase with disk induction test.

Clostridium difficile in the ICU: study of the incidence, recurrence, clinical characteristics and complications in a university hospital.

OBJECTIVE: Although several studies have established the association between antibiotics and Clostridium difficile infection (CDI), there is a lack of epidemiological studies on the incidence of CDI in European Intensive Care Units outside the context of infection outbreaks. The present study describes the incidence, patient characteristics, complications, and recurrence rates of CDI in a Spanish ICU. DESIGN: A retrospective study was carried out. SETTING: A clinical-surgical ICU with 34 beds, a tertiary referral hospital with 1400 beds. PATIENTS: All patients over 18 years of age admitted to the ICU from January 2010 to December 2011 with diarrhea for more than 48 h. INTERVENTIONS: None. STUDY VARIABLES: Underlying diseases, risk factors, fever, leukocyte count, complications, recurrence of infection. RESULTS: A total of 1936 adult patients were admitted. Seven patients acquired CDI (0.36%), representing an infection rate of 3.1 per 10,000 bed-days and a cumulative incidence rate of 3.6 in two years. The mean age was 61 years. Six patients showed some degree of immunosuppression. The mean APACHE II score at ICU admission was 17 (IQR 13-24). Severe sepsis was reported in 5 cases of CDI, three of which presented shock and multiorgan dysfunction. Four patients presented recurrence of CDI during hospitalization. ICU admission was prolonged for a mean of 24 days (SD 17.8) after diagnosis. CONCLUSIONS: Less than 1% of the patients admitted to a clinical-surgical ICU in a large teaching institution in Spain developed CDI. However, a high risk of recurrence/complications was associated with prolonged ICU stay.

Violacein improves vemurafenib response in melanoma spheroids.

Chemotherapy resistance is one of the main challenges in melanoma treatment. Violacein, a natural pigment produced by Chromobacterium violaceum, induces apoptosis in a variety of tumours, including melanoma. Here, we used BRAF-mutated melanoma spheroids to test the potential of violacein as a sensitizer of cellular viability and levels of the proteins p62 and fatty acid synthase (FASN). Importantly, violacein in combination with vemurafenib (ViVe) was able to interfere with spheroid survival at subtoxic concentrations. The results demonstrated that the ViVe protocol triggered cell death assessed by calcein and ethidium homodimer dyes. Accordingly, melanoma cells in 2D systems also showed a higher apoptosis rate when treated with ViVe. In the current study, we show evidence that ViVe downregulates crucial mediators like FASN, which partially explains how it acts as a sensitizer and ultimately improves the effectiveness of vemurafenib against melanoma cells.

New synthetic nano-immunotherapy (OncoTherad®) for non-muscle invasive bladder cancer: Its synthesis, characterization and anticancer property.

Bacillus Calmette-Guérin (BCG)-based intravesical immunotherapy has been applied as gold standard treatment for high-risk non-muscle invasive bladder cancer (NMIBC) for almost half a century. However, several patients with high-risk disease experience relapse, including those whose condition has worsened and who failed to respond to BCG. Non-significant therapeutic options have been developed for these at-risk patients, for many years. Immunotherapies have shown promising outcomes for bladder cancer treatment. Accordingly, our research group developed the OncoTherad® (MRB-CFI-1) immunotherapy, which has shown positive outcomes in NMIBC treatment. The aim of the current study is to describe, in details, the physicochemical features and potential action mechanisms of OncoTherad® nano-immunotherapy, based on toll-like receptor 4 (TLR4)-mediated interferon and on RANK/RANKL signaling pathways, in animal model with NMIBC. Based on the current findings, OncoTherad® nano-immunotherapy did not have genotoxic effect on the investigated model and did not show signs of limiting local and/or systemic toxicity at therapeutic doses. OncoTherad® nano-immunotherapy was more effective than the BCG treatment, since it reduced by 70% the malignancy rate. Furthermore, it was possible identifying an important action mechanism of OncoTherad®, which was based on the modulation of TLR4-mediated interferon and RANK/RANKL signaling pathways that, altogether, were essential to reduce malignancy rate. OncoTherad® mechanisms in these pathways helped preventing tumor recurrence.

The prevalence of interdigital erythrasma in southern region of Turkey.

BACKGROUND: Erythrasma is a skin infection which is caused by Corynebacterium minutissimum. Interdigital erythrasma is the most common form. OBJECTIVE: The aim of this study was to detect the frequency and risk factors of interdigital erythrasma in patients with clinically suspected tinea pedis. METHODS: This study was conducted between June and December 2010 and included 122 patients who had interdigital foot lesions. All patients were examined using a Wood's lamp. The smears were stained using Gram's method. Direct examination was performed using 20% potassium hydroxide. Sabouraud dextrose agar and brain heart infusion agar were used for cultures. Moreover, the demographical characteristics of patients, concomitant diseases and clinical findings were also recorded. Cases that were found to be positive on Wood's lamp examination and/or Gram staining/culture were considered as erythrasma. RESULTS: The rate of erythrasma was found to be 46.7%. The mean age was 43.6 years, and the disease was more prevalent in men. The most common clinical finding was desquamation. Mycological examination was found as positive in 40.35% of the patients. No growth was observed in bacteriological cultures. It was found that using only Wood's lamp examination or Gram staining resulted in 11 (9%) and 19 positive patients (15.6%), respectively, whereas using both Wood's lamp examination and Gram staining concurrently resulted in 27 positive patients (22.1%). CONCLUSION: Interdigital erythrasma is a commonly seen condition and can clinically mimic tinea pedis. A Wood's lamp is a good diagnostic tool, but Gram staining, particularly in those with a negative Wood's lamp result, may be a useful method.

OncoTherad® (MRB-CFI-1) nano-immunotherapy reduced tumoral progression in non-muscle invasive bladder cancer through activation of Toll-like signaling pathway.

The new modalities for treating patients with high-grade non-muscle invasive bladder cancer (HGNMIBC) for whom Bacillus Calmette-Guerin (BCG) has failed or is contraindicated are recently increasing due to the development of new drugs. Since NMIBC is sensitive to immunotherapy, Toll-like receptors (TLRs) agonist compounds may represent a potential antitumor therapeutic approach. Our research group developed a synthetic compound, with antitumor and immunological properties, called OncoTherad® (MRB-CFI-1). To evaluate the effects of OncoTherad® (MRB-CFI-1) and its compounds (P14-16 and CFI-1), thirty-six female C57Bl/6 J mice were divided into six groups (n = 6): Control, Cancer, Cancer + BCG (40 mg), Cancer + OncoTherad® (20 mg/mL), Cancer + P14-16 (20 mg/mL) and Cancer + CFI-1 (20 mg/mL). NMIBC was chemically induced (N-ethyl-N-nitrosourea 50 mg/mL) and the treatments were followed for six weeks. The bladder was collected and routinely processed for immunohistochemical analyses of the Toll-Like receptors signaling pathway (TLR2, TLR4, MyD88, IRF-3, IKK-α, NF-kB, TNF-α, TRIF, IFN-γ, IL-6). The results obtained showed that the tumor progression was 100 % reduced on OncoTherad® (MRB-CFI-1) treated animals. Immunohistochemical analysis demonstrated that while the conventional BCG treatment stimulated the canonic pathway, OncoTherad® (MRB-CFI-1) stimulated the non-canonical pathway (increasing expression of TLR4, TRIF, IRF, and IFNγ). OncoTherad® (MRB-CFI-1) could be considered a promising therapy in the treatment of NMIBC.

Descriptive study of a telemedicine program in palpebral pathology for primary care (tele-eyelid).

BACKGROUND AND OBJECTIVE: In 2013 we implemented an asynchronous telemedicine circuit for the diagnosis of eyelid diseases (tele-eyelid), connecting the outpatient primary healthcare with the hospital's specialists. The purpose of this study is to assess the use of telemedicine in the diagnosis of eyelid diseases by primary care teams, to evaluate its usefulness and to analyse the epidemiology of the pathological conditions referred to the tertiary level hospital, as well as the need for surgery. MATERIALS AND METHODS: This study was carried out in the Spanish public health system, in a mainly rural area assisted by the Institut Català de la Salut (ICS) and Althaia Xarxa Assistencial Universitària de Manresa. This is a retrospective, descriptive analysis of the telematic consultations undertaken between 2013 and 2019. The consultations between 2018 and 2019 underwent a further descriptive retrospective-prospective analysis to assess the conditions referred to the hospital. RESULTS: Unnecessary referrals were avoided in 72% of telematic consultations. More than 50% of primary care practitioners used tele-eyelid. Up to 68% of the referrals were due to eyelid tumours, 50% needed surgery and 18%, a biopsy. Moreover, we found a high reliability between telematic and face-to-face diagnosis. CONCLUSIONS: Teleophthalmology applied to eyelid pathology is a useful tool to improve access to specialized care and helps solving pathological conditions. It avoids unnecessary consultations and increases efficiency, both in primary and hospital care.

Nanostructured polymer and lipid carriers for sunscreen. Biological effects and skin permeation.

The interest in developing new sunscreens is increasing due to the harmful effects of UV radiation on the skin, such as erythema, accelerated skin ageing (photoageing) and the induction of skin cancer. However, many molecular sunscreens penetrate into the skin causing photoallergies, phototoxic reactions and skin irritation. Thus, the aim of this work was the preparation and characterization of polymeric and solid lipid nanoparticles to act carriers of benzophenone-3 (BZ3), aiming to improve the safety of sunscreen products by increasing the sun protection factor (SPF), decreasing BZ3 skin penetration and decreasing BZ3 concentration in sunscreen formulation. BZ3 was encapsulated in poly(epsilon-caprolactone) (PCL) nanoparticles by the nanoprecipitation method and in solid lipid nanoparticles (SLN) by the hot high pressure homogenization method. The particles were stable for 40 days. The BZ3 encapsulated in PCL nanoparticles was released faster than BZ3 encapsulated in SLN. The sun protection factor increased when BZ3 was encapsulated in both nanostructures. However, BZ3 encapsulated in PCL nanoparticles decreased its skin permeation more than SLN-BZ3. Furthermore, BZ3 encapsulated in SLN did not exhibit cytotoxic or phototoxic effects in human keratinocytes (HaCaT cells) and BABL/c 3T3 fibroblasts, whereas PCL nanoparticles with BZ3 showed phototoxic potential in HaCaT cells. Nevertheless, BZ3 free and encapsulated in PCL nanoparticles or in SLN did not show allergic reactions in mice. Our results suggest that these nanostructures are interesting carriers for sunscreen.

Design of magnetic hybrid nanostructured lipid carriers containing 1,8-cineole as delivery systems for anticancer drugs: Physicochemical and cytotoxic studies.

The development of versatile carriers to deliver chemotherapeutic agents to specific targets with establishing drug release kinetics and minimum undesirable side effects is becoming a promising relevant tool in the medical field. Magnetic hybrid nanostructured lipid carriers (NLC) were prepared by incorporation of 1,8-cineole (CN, a monoterpene with antiproliferative properties) and maghemite nanoparticles (MNPs) into a hybrid matrix composed of myristyl myristate coated with chitosan. Hybrid NLC characterized by DLS and TEM confirmed the presence of positively charged spherical nanoparticles of around 250 nm diameter and +10.2 mV of Z-potential. CN encapsulation into the lipid core was greater than 75 % and effectively released in 24 h. Modification of the crystalline structure of nanoparticles after incorporation of CN and MNPs was observed by XRD, DSC, and TGA analyses. Superparamagnetic NLC behavior was verified by recording the magnetization using a vibrating scanning magnetometer. NLC resulted in more cytotoxic than free CN in HepG2 and A549 cell lines. Particularly, viability inhibition of HepG2 and A549 cells was increased from 35 % to 55 % and from 38 % to 61 %, respectively, when 8 mM CN was incorporated into the lipid NPs at 24 h. Green fluorescent-labeled NLC with DIOC18 showed an enhanced cellular uptake with chitosan-coated NLC. Besides, no cytotoxicity of the formulations in normal WI-38 cells was observed, suggesting that the developed hybrid NLC system is a safe and good potential candidate for the selective delivery and potentiation of anticancer drugs.

Development of nanostructured silver vanadates decorated with silver nanoparticles as a novel antibacterial agent.

In this work we report the synthesis, characterization and application of silver vanadate nanowires decorated with silver nanoparticles as a novel antibacterial agent. These hybrid materials were synthesized by a precipitation reaction of ammonium vanadate and silver nitrate followed by hydrothermal treatment. The silver vanadate nanowires have lengths of the order of microns and diameters around 60 nm. The silver nanoparticles decorating the nanowires present a diameter distribution varying from 1 to 20 nm. The influence of the pH of the reaction medium on the chemical structure and morphology of silver vanadates was studied and we found that synthesis performed at pH 5.5-6.0 led to silver vanadate nanowires with a higher morphological yield. The antimicrobial activity of these materials was evaluated against three strains of Staphylococcus aureus and very promising results were found. The minimum growth inhibiting concentration value against a MRSA strain was found to be ten folds lower than for the antibiotic oxacillin.

The effects of dimensionality on electrochemical sensors based on carbon nanotubes and metallic nanowires.

Electrochemistry based unidimensional nanoelectrodes, such as carbon nanotubes and metallic nanowires, is now a reality that is leaving the academy and reaching a broad range of high-tech companies around the world. The intrinsic properties of electron-electron interactions, sizes and geometries significantly differentiate those nanostructures from conventional macroelectrodes or electrodes with sizes in a millimetric range. The inherent properties as well as the configuration of carbon nanotubes and metallic nanowires as electrochemical sensors are the main focus of this review. Applications and some future trends on developing of a clear pathway for assembly and integration of these into functional materials, are summarized as well.

Antimicrobial activity of the extracts and physalin D from Physalis alkekengi and evaluation of antioxidant potential of physalin D.

Physalis alkekengi L. (Solanaceae) is a popular plant in traditional European and Chinese folk medicine, and it has been reported to have many ethnopharmacological properties including antifungal, anti-cough, anti-inflammatory, analgesic, and febricide activities. Some active components from Physalis species have been investigated. However, no antimicrobial activity studies on extracts and physalins of P. alkekengi have been carried out. In this study, we attempted to identify the possible antimicrobial activities of the methanol extract from aerial parts of P. alkekengi and the dichloromethane extract from calyces of the plant. The extracts were tested against five Gram-positive and five Gram-negative bacteria and five Candida species by using disk diffusion and broth microdilution methods. The extracts were fractionated to isolate physalins using chromatographic techniques, and physalin D was isolated from the extracts. The structure of the compound was elucidated on the basis of (1)H-NMR spectroscopic study, and confirmed by comparison with a reference sample and literature data. Results indicated that all the extracts and physalin D were characterized by antibacterial action, especially against Gram-positive bacteria, with MIC values between 32 and 128 microg/mL. The methanol extract had moderate activity against fungi at MICs ranging from 128 to 512 microg/mL, but the dichloromethane extract and physalin D had low activity against fungi at MICs ranging from 256 to 512 microg/mL. Additionally, the antioxidant activity of physalin D was evaluated by qualitative DPPH (1,1-diphenyl-2-picrylhydrazyl) radical and TBA (thiobarbituric acid) assays. Physalin D showed low antioxidant activity with an IC(50) value of >or= 10 +/- 2.1.

Evaluation of oral streptococci in saliva of children with severe Early Childhood Caries and caries-free.

AIM: Oral streptococci were found to be associated with Early Childhood Caries. The aim of this study was to evaluate the 6 different bacteria in the streptococcus group in the saliva of children with severe early childhood caries (S-ECC) by polymerase chain reaction (PCR). MATERIALS AND METHODS: A total of 60 children between 3 and 6 years of age were divided into two groups: children with S-ECC (Group S-ECC; n=30) and children who were caries-free (Group CF; n=30), according to the dmft and dmfs indices. Unstimulated saliva was collected from all participants for the detection of streptococcal group bacteria, including: Streptococcus mutans, Streptococcus oralis, Streptococcus sanguinis, Streptococcus gordonii, Streptococcus salivarius, and Streptococcus sobrinus, using PCR-restriction fragment length polymorphism (RFLP) of amplified 16S rRNA gene. The data were analysed using SPSS software. RESULTS: The prevalence of S. oralis was significantly higher in the S-ECC group compared to the CF group (p<0.05). However, the frequencies of S. mutans, S. sanguinis, S. gordonii, S. salivarius, and S. sobrinus were similar between the two groups (p>0.05). The amount of streptococci colonies was higher in the S-ECC group compared to the CF group (p<0.05). CONCLUSION: S. mutans or S. sobrinus alone may not be the only indicators for high risk of caries, but the prevalence of S. oralis in saliva may be a risk factor for increased caries activity in S-ECC.

Silver nanoparticles: Toxicity in model organisms as an overview of its hazard for human health and the environment.

Silver nanoparticles (AgNPs) have attracted remarkable attention due to their powerful antimicrobial action as well as their particular physicochemical properties. This has led to their application in a wide variety of products with promising results. However, their interaction with the environment and toxicity in live terrestrial or aquatic organisms is still a matter of intense debate. More detailed knowledge is still required about the toxicity of AgNPs, their possible uptake mechanisms and their adverse effects in live organisms. Several studies have reported the interactions and potential negative effects of AgNPs in different organisms. In this review, we report and discuss the current state of the art and perspectives for the impact of AgNPs on different organisms present in the environment. Recent progress in interpreting uptake, translocation and accumulation mechanisms in different organisms and/or living animals are discussed, as well as the toxicity of AgNPs and possible tolerance mechanisms in live organisms to cope with their deleterious effects. Finally, we discuss the challenges of accurate physicochemical characterization of AgNPs and their ecotoxicity in environmentally realistic conditions such as soil and water media.

Biodegradation and ecotoxicological assessment of pectin production wastewater.

The chemical composition of pectin production wastewater and its toxicity during biological treatment were investigated. Samples of wastewater from different steps of a pectin production wastewater biological treatment plant were investigated including the influent of the treatment (1), after denitrification tank (2), after anaerobic treatment (3) and final effluent (4). The conventional physicochemical characteristics of samples did not indicate wastewater toxicity. However, toxicity assessments carried out on Vibrio fischeri and Scenedesmus subspicatus indicated low EC50 values. The fractionation of the samples using an XAD resin showed that the toxicity was associated with the organic matter. Wastewater apparent molecular mass distributions were 14.3, 25.0, 24.4 and 29.6 kDa for samples 1-4, respectively. Finally, characteristics of the sample by pyrolisis-gas chromatography-mass spectrometry (Py-CG-MS) demonstrated its polyphenolic nature and a 23% increase in the levels of such compounds after the first biological treatment step.

Chronic dacryocystitis caused by sarcoidosis. / Dacriocistitis crónica causada por sarcoidosis.

Sarcoidosis of lacrimal sac is a very uncommon disease. When the patient has symptoms of lacrimal sac diseases, it is necessary to take into account the histological evaluation, to demonstrate the involvement of the lacrimal sac by this pathology. In patients with sarcoidosis history, the treatment is surgical and it is suggested to do an external dacryocystorhinostomy, in order to take a biopsy, to get a diagnosis and to establish the incidence of this pathology. The case is presented of a 44 year-old female, with a history of sarcoidosis lymph nodes, who presented with a right chronic dacryocystitis of one year onset.