Mycetoma caused by Fusarium solani with osteolytic lesions on the hand: case report

Eumycetoma is a mycotic disease caused by saprophytic soil fungi that are usually inoculated through minor injuries. A case of mycetoma in a Brazilian farmer aged71 years is reported. This patient presented erythema and edema on the dorsal surface of the left hand with multiple crusted and cicatricial lesions. No macroscopic grains were observed. The histopathological findings showed grains consisted of numerous hyphae which stained well with Gomori-Grocott method. This material obtained by cutaneous biopsy was submitted to culture on Sabouraud’s medium and the colonies were identified as Fusarium solani. The radiological studies revealed bone osteolytic lesions and the ultrasound showed pseudocysts and fistulae at the site of this infection. The patient was treated with oral ketoconazole with a good clinical response. This revised version was published online in June 2006 with corrections to the Cover Date.     

A clinico-pathological study of actinomycotic mycetomas caused by Actinomadura madurae and Actinomadura pelletierii

Twenty seven cases of actionomycotic mycetoma caused either by Actinomadura madurae or Actinomadura pelletierii have been described. Infection by A. madurae has been more common than A. pelletierii. Left foot in A. madurae and right foot in A. pelletierii infections were involved more commonly in adult males, whereas right foot of the females was frequently affected in A. madurae infection. Large, soft, white grains in A. madurae and small, firm, red grains in A. pelletierii were consistantly seen. Deep hematoxylin stained grains with scalloped margin and prominent eosinophilic club in A. madurae and such deep stained grains with smooth margin and horizontal cracks appearing as protions of a spherical mass in A. pelletierii were diagnostic. Large numbers of plasma cells and Russel bodies were also characteristic of A. madurae infection. Both the grains were stainable with Von Kossa method for calcium. Bone changes were similar in both the infections. Oral tetracycline produced soft tissue and bone resolution to almost normalcy in those who regularly consumed the drug any time from 2 to 6 years. Mild glucose intolerance, facial hyperpigmentation and urticaria were the side effects observed in a few. Two patients developed cataract following tetracycline therapy. The value of medical therapy with oral tetracycline in Actionomadura mycetomas is emphasized.     

A sensitive method for detection of proteins in polyacrylamide gels and on protein blots using acidic henna leaf extract

Summary In the present communication we describe a process for the preparation of an acidic henna leaf extract useful as a general protein stain for both polyacrylamide gels and protein blots. The staining is reversible by changing its pH and does not require protein fixation or destaining steps. This staining procedure is simpler and also, its sensitivity is comparable with the two commonly used organic stains i.e. Coomassie Blue and Amido Black. Under optimum conditions the protein detection limits of acidic henna leaf extract varied from 50 ng to 100 ng.     

A case of eumycetoma due to Madurella grisea in northern Brazil

A case of mycetoma caused by the black fungus Madurella grisea in northern Brazil is reported. The lesion was located on the patient’s right foot without bone involvement. Clinical samples were collected by opening the fistules with a scalp and the grains removed for microbiological and histopathological analyses. Although mycetoma caused by M. grisea has been previously reported in Brazil, this is the first time that of this fungus has been identify in this particular region of the country.     

Hand Mycetoma: The Mycetoma Research Centre Experience and Literature Review

Mycetoma is a devastating, neglected tropical disease characterised by extensive tissue involvement resulting in destruction, deformities and disabilities in the affected patients. The hand is commonly affected by mycetoma thus compromises its functionality and hinder the patient’s daily activities of living. In this communication, we report on 533 patients with hand mycetoma managed over a period of 24 years at the Mycetoma Research Centre, University of Khartoum, Khartoum, Sudan. Eumycetoma was the commonest type of mycetoma (83.3%) encountered. Males were predominately affected (69.2%) with a sex ratio of 2.2:1. The majority of the patients (84%) were young adult below the age of 40 years old at presentation. The generality of patients (86.4%) were from the Sudan mycetoma belt. Children and adolescents (28.1%), farmers (18.2%) and workers (17.4%) were more frequently affected. The majority of patients (67.4%) had disease duration of less than 5 years at presentation. The study, did not document significant history of local trauma, familial tendency, concomitant medical diseases or other predisposing cause for mycetoma in this population. Pain (23.1%) was not a disease feature in this series and 52% of patients had past surgery for mycetoma and recurrence. The right hand was affected most (60.4%), and 64% of them had small lesion at presentation. Conventional x-ray was only helpful in patients with advanced disease and the MRI accurately determined the disease extension. Cytological smears, surgical biopsies histopathological examination and grains culture were the principal diagnostic tools for causative organisms’ identification. In the present series it was difficult to determine the treatment outcome due to high patients follow up dropout.     

用花粉粒染色法标记虫媒植物的花粉流

花粉作为植物有性生殖过程中的雄性供体,其释放后的去向和散布范围直接影响植物的交配模式和居群遗传结构,但花粉的运动肉眼不易观察,因此标记花粉运动的方法一直是传粉生物学中的关键技术［１,２］。尽管已有多种分子标记的技术被用来研究花粉的运动,包括同工酶［３...     

Is the surface layer from hazelnut pollen,which is precipitated by Cuprolinic blue,an effective antigen in hay-fever patients?

Summary In electron micrographs it could be shown that hazelnut (Corylus avellana) pollen grains are covered on their surface by a diffusible 10 nm thick lamellar layer. On pollen surface as well as in pollen extract this layer could be precipitated and stained by the polycationic dye Cuprolinic blue. By subsequent application of both immunogold labeling with serum from a hay-fever patient allergic to tree pollen grains and histochemical detection with Cuprolinic blue this pollen surface layer proved to be an effective antigen.     

Lawsone,a 2-hydroxy-1,4-naphthoquinone from Lawsonia inermis (henna), produces mitochondrial dysfunctions and triggers mitophagy in Saccharomyces cerevisiae

Molecular Biology Reports - Lawsone is a natural naphthoquinone present in the henna leaf extract with several cytotoxic activities and used as precursor for synthesis of various pharmaceutical...     

Vital and Calcofluor staining ofCosmarium and its protoplasts

Summary Intact cells, protoplasts, primary and secondary walls ofCosmarium species were stained with 13 vital stains and with Calcofluor. The intracellular contents of viable cells and protoplasts were not stained with any of the test dyes although the primary and secondary walls often were stained. Protoplasts disintegrated after short periods in acid stains but did survive for up to 24 hours in several dilute basic dyes. Once the protoplast membrane was damaged, the nuclei and cytoplasm were brightly stained with most dyes, whereas the vacuoles remained unstained. Calcofluor was particularly useful in determining the porous structure and pattern of the primary walls.     

Mycetoma in the Sudan: An Update from the Mycetoma Research Centre,University of Khartoum,Sudan

This communication reports on the Mycetoma Research Centre of the University of Khartoum, Sudan experience on 6,792 patients seen during the period 1991–2014.The patients were predominately young (64% under 30 years old) males (76%). The majority (68%) were from the Sudan mycetoma belt and 28% were students. Madurella mycetomatis eumycetoma was the most common type (70%). In 66% of the patients the duration of the disease was less than five years, and 81% gave a history of sinuses discharging mostly black grains (78%). History of trauma at the mycetoma site was reported in 20%. Local pain was reported in 27% of the patients, and only 12% had a family history of mycetoma. The study showed that 57% of the patients had previous surgical excisions and recurrence, and only 4% received previous medical treatment for mycetoma. Other concomitant medical diseases were reported in 4% of the patients. The foot (76%) and hand (8%) were the most commonly affected sites. Less frequently affected sites were the leg and knee (7%), thigh (2%), buttock (2%) and arm and forearm (1%). Rare sites included the chest wall, head and neck, back, abdominal wall, perineum, oral cavity, tongue and eye. Multiple sites mycetoma was recorded in 135 (2%) of cases. At presentation, 37% of patients had massive lesions, 79% had sinuses, 8% had local hyper-hydrosis at the mycetoma lesion, 11% had regional lymphadenopathy, while 6% had dilated tortuous veins proximal to the mycetoma lesions. The diagnosis of mycetoma was established by combined imaging techniques and cytological, histopathological, serological tests and grain culture. Patients with actinomycetoma received a combination of antimicrobial agents, while eumycetoma patients received antifungal agents combined with various surgical excisions. Surgical excisions in the form of wide local excision, debridement or amputation were done in 807 patients, and of them 248 patients (30.7%) had postoperative recurrence. Different types of amputations were done in 120 patients (1.7%).     

Active Matrix Metalloprotease-9 Is Associated with the Collagen Capsule Surrounding the Madurella mycetomatis Grain in Mycetoma

Madurella mycetomatis is the main causative organism of eumycetoma, a persistent, progressive granulomatous infection. After subcutaneous inoculation M. mycetomatis organizes itself in grains inside a granuloma with excessive collagen accumulation surrounding it. This could be contributing to treatment failure towards currently used antifungal agents. Due to their pivotal role in tissue remodelling, matrix metalloproteinases-2 (MMP-2) and 9 (MMP-9) or tissue inhibitor of metalloproteinases (TIMP) might be involved in this process. Local MMP-2 and MMP-9 expression was assessed by immunohistochemistry while absolute serum levels of these enzymes were determined in mycetoma patients and healthy controls by performing ELISAs. The presence of active MMP was determined by gelatin zymography. We found that both MMP-2 and MMP-9 are expressed in the mycetoma lesion, but the absolute MMP-2, -9, and TIMP-1 serum levels did not significantly differ between patients and controls. However, active MMP-9 was found in sera of 36% of M. mycetomatis infected subjects, whereas this active form was absent in sera of controls (P<0.0001). MMP-2, MMP-9, and TIMP-1 polymorphisms in mycetoma patients and healthy controls were determined through PCR-RFLP or sequencing. A higher T allele frequency in TIMP-1 (+372) SNP was observed in male M. mycetomatis mycetoma patients compared to controls. The presence of active MMP-9 in mycetoma patients suggest that MMP-9 is activated or synthesized by inflammatory cells upon M. mycetomatis infection. Inhibiting MMP-9 activity with doxycycline could prevent collagen accumulation in mycetoma, which in its turn might make the fungus more accessible to antifungal agents.     

Novel fluorescent dyes based on oligopropylamines for the in vivo staining of eukaryotic unicellular algae

Weakly basic fluorescent dyes are used to visualize organelles within live cells due to their affinity to acidic subcellular organelles. In particular, they are used to stain the silica deposited in the silica deposition vesicles (SDVs) of diatoms during the course of their frustule synthesis. This study involved the synthesis of fluorescent dyes derived from oligopropylamines, compounds similar to those found in diatoms. The dyes were obtained by reacting oligopropylamines with 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole. The reaction was realized using methylated oligopropylamines with two or three nitrogen atoms and yielded two novel fluorescent dyes: NBD-N2 and NBD-N3. The dyes appeared to be highly efficient in the in vivo staining of growing siliceous frustules of diatoms at concentrations at least 10 times lower than those required for staining with HCK-123. NBD-N3 also efficiently stained other subcellular vesicles of eukaryotic unicellular algae. NBD-N2 stained only growing diatom frustules, whereas NBD-N3 also stained various subcellular organelles of different eukaryotic unicellular algae. NBD-N2 and NBD-N3 were not removed from stained diatom frustules by drastic treatments with H₂SO₄ and H₂O₂. Fluorescent silica can also be obtained by its chemical precipitation in the presence of NBD-N2 and NBD-N3.     

Hypersensitivity reactions due to black henna tattoos and their components: are the clinical pictures related to the immune pathomechanism?

Hypersensitivity to para-phenylenediamine (PPD) and related compounds induced by temporary black henna tattoos has become a serious health problem worldwide. Different patterns of sensitization with various clinical aspects are described in literature due to PPD associated to henna tattoo and these manifestations are likely correlated with the immunological and dermatological pathomechanisms involved. Henna is the Persian name of the plant Lawsonia inermis, Fam. Lythraceae. It is a woody shrub that grow in regions of North Africa, South Asia, India and Sri Lanka. Nowadays it is rather frequent to see temporary “tattoos” performed with henna. To make tattoos darker and long-lasting PPD has been associated to henna in tattoo drawings mixtures, so obtaining “black henna”. In these years there has been a rise of contact sensitization to PPD and in medical literature an increased number of cases have been reported on temporary henna tattoo application. Here we review the various clinical patterns related to PPD and henna tattoo, to investigate the possible link between clinic-morphological pictures and the immunological response to PPD and henna. The literature underlines that different clinical manifestations are related to black henna containing PPD, and its derivative products may cause delayed-type as well as immediate-type reactions. Further studies are needed to investigate the relationship between clinical and morphological aspects of PPD contact dermatitis and the T cell subsets predominance.     

Multi-photon microscopy of cell types in the viable taste disk of the frog

The morphology of viable taste disks of the frog was explored with multi-photon microscopy. In order to identify single sensory or supporting cells within the tissue, we searched for fluorescent dyes that stained subsets of the cell population or possibly cell types. Some cell types indeed stained preferentially with certain fluorescent dyes. A subset of glia-like cells (type Ic) stained with BCECF, a H⁺-sensitive dye, and indo-1, a Ca²⁺-sensitive dye, both presented in the membrane-permeant ester form. BCECF-ester also stained the dendrites of type III receptor cells, but indo-1 ester did not. Receptor cells of type II stained with MQAE, a positively charged Cl^–-sensitive dye. A subset of type II cells accumulated amiloride, a positively charged fluorescent diuretic. Certain supporting cells, i.e., wing cells (type Ib) and glia-like cells (type Ic), were labeled by negatively charged dyes, e.g., calcium green-1 dextran. Mucus cells (type Ia) were stained with only two of the 19 dyes examined, and Merkel-like basal cells (type IV) were stained only with a membrane-labeling voltage-sensitive dye, presumably by endocytosis. No dye was found which would stain all types of cells or all receptor cells. This finding reveals a potential problem for future functional imaging aiming at population responses, as the responses of unstained cells then would remain unobserved. Specificity of dyes with respect to cell types was sufficient to identify supporting cells and receptor cells. Cell shape could then be reconstructed, using optical slicing and rendering techniques. Thus populations of dye-loaded elongated cells, especially types Ic, II and III, could for the first time be visualized in three dimensions.This work was supported by the Deutsche Forschungsgemeinschaft (SFB 530, project B2)     

Is the surface layer from hazelnut pollen, which is precipitated by Cuprolinic blue, an effective antigen in hay-fever patients?

In electron micrographs it could be shown that hazelnut (Corylus avellana) pollen grains are covered on their surface by a diffusible 10 nm thick lamellar layer. On pollen surface as well as in pollen extract this layer could be precipitated and stained by the polycationic dye Cuprolinic blue. By subsequent application of both immunogold labeling with serum from a hay-fever patient allergic to tree pollen grains and histochemical detection with Cuprolinic blue this pollen surface layer proved to be an effective antigen.     

Phototoxicity of the fluorescent membrane dyes PKH2 and PKH26 on the human hematopoietic KG1a progenitor cell line.

BACKGROUND: The phototoxic effects of the well-known fluorescent membrane dyes PKH2 and PKH26 have been unknown, although their use in cell tracking experiments has increased dramatically. To eliminate the phototoxicity-induced alteration in cell function and morphology, it is essential to examine the suspicious phototoxicity of these dyes. METHODS: Chemical and phototoxic effects of PKH dyes on the human hematopoietic KG1a cell line were examined. To minimize phototoxicity in long-term cell tracking experiments lasting up to 18 h with a fluorescence microscope system, time-lapse monitoring with different time intervals and exposure times was introduced. RESULTS: There were no significant effects of the two PKH dyes on cell viability and growth when using dye concentrations up to 5 microM. However, when stained cells were exposed to excitation light, cell viability decreased dramatically, showing the phototoxicity of the PKH dyes. More than 60% of cells stained with 5 microM PKH26 died after 5 min of continuous light exposure. The phototoxic effect was more extensive in cells stained with higher concentrations of the dyes. CONCLUSIONS: We present guidelines for the optimal use of these dyes by using a defined hardware configuration.     

Chromosome banding: specification of structural features of dyes giving rise to G-banding

Summary Metaphase chromosomes were stained in a routine G-banding procedure with 39 basic dyes of varied structures substituted for the Giemsa stain. Staining outcomes were categorized as: iverstained, differentially stained, trivially or unstained. Certain structural features of the dyes were described numerically, namely, largest conjugated fragment (LCF), conjugated bond number (CBN) and cationic weight. The staining, outcomes were compared to these numerical structural parameters, and structure-staining correlations sought. Dyes with large conjugated systems (and high LCF values) were seen to be overstained; dyes with low LCF values were often non-staining. At intermediate LCF values, the more hydrophobic dyes (with high Hansch values) stained differentially; the more hydrophilic dyes failed to stain. Expressed numerically, 89% of the dyes with the following characteristics stained differentially: 30LCF10; Hansch >–5.0. It was concluded that contributions to dye-chromosome affinity included coulombic forces and van der Waals attractions and that the selectivity of G-banding was largely due to hydrophobic bonding. Induction of bands could be due to the loss of hydrophilic, histones, amplifying underlying variations in the hydrophobic-hydrophilic character of the chromosome structure. Relatively hydrophobic sites include AT-rich DNA and disulphide-rich proteins.The effects on Romanowsky G-banding of chemically modifying chromosomes were in keeping with this model. Overstaining resulted from formation of either hydrophobic or conjugated derivatives or both, whereas trivial or non-staining arose from the formation of hydrophilic derivatives. Intriguingly, the efficacy of the dyes used for Q-banding also correlated positively with their hydrophobic character.     

Mature Cereal Grain Endosperm: Rapid Glass Knife Sectioning for Examination of Proteins

A simple, rapid procedure was developed by which the quality of endosperm protein in cereal grains could be evaluated by microscopic observation of the subcellular protein structure. Kernels with vitreous endosperm were sectioned without pretreatment at 3-4 μ with a glass knife. Floury endosperm tissues were fixed in 10% glutaraldehyde, pH 7.4, for 16 hr at 5 C, and boiled to gelatinize the starch. After drying, this tissue was sectioned as for vitreous endosperm. Sections were mounted on gelatin-coated slides and destarched with a-amylase. To identify sites of prolamine, alcohol-soluble protein was extracted for 1 hr at about 70 C with 80% ethanol. The subcellular proteins were stained with either iodine vapor or various organic dyes. Proteins were differentiated by a combination of staining and mounting in selected high refractive index liquids.     

A Call to Action for Mycetoma

Purpose of Review

Here, we discuss the current needs and priorities for mycetoma control and prevention, highlight lessons learned from leprosy and podoconiosis, and motivate an urgent need to accelerate progress toward reducing the burden of mycetoma in endemic areas.

Recent Findings

In 2015, the World Health Assembly (WHA) added mycetoma, a progressively debilitating disease caused by fungi and bacteria, to the World Health Organization (WHO) list of priority neglected tropical diseases (NTDs). Designation of other diseases as NTDs has raised awareness, enabled global partnerships, and advanced the capacity to combat disease through integrated programming. Although key mycetoma etiologic agents have been identified, many questions remain and mycetoma may similarly benefit from NTD designation.

Summary

In collaboration with experts at WHO and elsewhere, we formed a global mycetoma working group to connect partners from a variety of sectors and specialties. We envision that this group will evolve into a formalized partnership that can prioritize strategic planning, advocacy, and research needs, identify funding sources, and coordinate activities related to mycetoma and other NTDs affecting the skin. The experiences gained from other NTDs can help to guide the global mycetoma working group’s activities to better address the goals set forth in the WHA resolution.

     

Mechanism of connective tissue techniques I. The effect of dye concentration and staining time on anionic dye procedures

Summary Anionic dye connective tissue procedures were performed by staining for 5 min and 24 h with (a) 0.00018m and 0.0018m solutions of 28 dyes, and 0.018m solutions of 21 dyes in saturated picric acid (SPA), and (b) 0.0018m and 0.018m solutions of 20 dyes in 1% (w/v) phosphomolybdic acid (PMA). The staining obtained with dyes in SPA was classified as selective (no cytoplasmic staining), moderately selective (traces of cytoplasmic staining) and non-selective (all other staining patterns). The staining of collagen and cytoplasm with dyes in PMA was separately classified on a scale of 1–5 (1 = no staining, 5 = maximum staining). The selectivity of the staining obtained with SPA with solutions of dyes at concentrations of 0.00018m and 0.0018m, and both staining times, was correlated (p < 0.001) with an empirical sulphonic acid constant (SAC) defined as the (number of dye sulphonic acid groups/dye molecular weight) × 10³. Correlation with molecular weight was poor and was significant only when staining was performed with 0.00018m dye solutions for 24 h. The dyes were divisible into three groups: group 1 (selectivity independent, or almost independent of staining time), group 2 (selective to moderately selective when staining was performed for 5 min), and group 3 (non-selective). The SAC of the group 1 dyes differed significantly from those of the group 2 and 3 dyes. Selectivity was essentially lost at dye concentrations of 0.018m. The staining with acidic dyes (no amines or substituted amines) in PMA differed significantly (p < 0.001) from that obtained with amphoteric dyes (containing basic substituents). In general, acidic dyes stained cytoplasm. Amphoteric dyes with the exception of indigocarmine stained collagen. However, most of these dyes also stained cytoplasm. In contrast to the results obtained with dyes in SPA, selectivity correlated strongly with molecular weight and only poorly with the SAC. Staining time and dye concentration affected selectivity only when the acidic dyes were used for 5 min at concentrations of 0.0018m and 0.018m. The data obtained do not permit a clear distinction between the rate control and chemical affinity models for the mechanism of staining with anionic dyes. However, it seems possible that different groups of dyes stain by different mechanisms. Part of this work was performed by M.I., S.N., M.J. and L.M. in partial fulfilment of the requirements for the completion of Pathology 438. A partial account of this work was presented at the annual convention of the British Columbia Society of Medical Technology, Victoria, British Columbia, October 1991.