The chemotactic activity of Candida albicans proteinase toward human peripheral leuko- cytes was examined by a modified Boyden chamber method.It was found that C.albicans proteinase in- duce dose-dependent chemotactic activity for neutrophils with maximal activity at 500 nmol/L(P

Objective To report a case of black-dot ringworm caused by Trichophyton tonsurans in a 3-year-old girl. Methods Lesional hair was obtained from the patient and subjected to direct microscopic examination as well as culture. Subsequently, the isolate underwent morphological, biochemical and molecular biology identification. The extracellular enzymatic activity of the isolate was analyzed. Results Microscopy revealed that the hair shaft was filled with fungal spores. Typical colony of the isolate was grayish-white with downy appearance. Slide culture showed centipede-like, lateral, rod-shaped microconidia. Urease test was positive. The amplification of ribosomal DNA (rDNA) ITS domains by PCR produced a 687 bp-sized fragment which had a 100% homology with the sequences of several Trichophyton tonsurans strains in the GenBank database. The extracellular enzymatic activity analysis showed an increase in the activity of alkaline phos-phatase, acid phosphatase, esterase (C4), β-glucosidase, leucine arylamidase, N-acetyl-β-glucosaminidase and a-mannosidase. Conclusions The pathogenic fungus is identified as Trichophyton tonsurans based on morphological and biochemical features as well as sequence of the ITS region of rDNA, and the child was diagnosed with black-dot ringworm.

Objective To isolate and observe Pityrosporum yeasts from a patient with onychomycosis. Methods The involved nail specimens were investigated by means of culture, pathological and scanning electron microscopic examination and 20% KOH preparation. Results Physical examination showed that each finger and toe nail appeared brownish black, rough and thick, some of the fore part of the nail plate detached from the nail bed. Fingernail specimen's culture results showed that Trichophyton rubrum grew on Sabouraud's dextrose agar and Pityrosporum ovale grew on the medium containing rapeseed oil. The pathological examination revealed P.ovale yeast involvement in the fissure of the nail plate. Under the scanning electron microscopy, a lot of P.ovale yeasts with characteristic collarette structure inserted in the nail tissue was noticed. In the 20% KOH preparations of nail incubated at 56℃for 1h and stained with Quink Parker ink, spores and hyphae were identified morphologically with P.ovale and T.rubrum respectively. The patient received intermittent pulse therapy with itraconazole, the color of the nails became much brighter 1 to 2 months after the fourth cycle of therapy, but no further improvement was observed afterwards. P.ovale and T.rubrum grew again 6 months after treatment when the clippings of the fingernail were cultured. Conclusion This is the first document of onychomycosis related with P.ovale in China.

Objective To study the morphological,biologic response of cultured human keratin ocytes to Candida albicans and the molecular mechanism.Methods ①A yeast form of C.albicans was added t o a monolayer of keratinocytes.The adherent organisms were observed by scanning and transmission electron mi-croscope at different incubation periods.Boiled C.albicans and latex beads of a similar size(3.2?m)were also inoculated for comparison.②The effect of supernatant from the co-culture of keratinocytes with inta ct C.albicans on keratinocyte growth was measured by the fluorescence intensity and cell count.③Keratinocytes were incubated with C.albicans or latex beads,and the level of cytok ine in the keratinocytes and supernatant were measured with enzyme -linked im munosorbent assay kits.Results①The number of adherent C.albicans increased with the lapse of time,whi le boiled C.albicans did not adhere at all.Many latex bead s adhered to the keratinocytes,and were then easily phagocytised.Fibril -like structu res stretched from the keratinocyte s adhered to the organisms or latex beads.②The conditioned medium of 50%concen tration significantly promote cell growth,while that with boiled C.albicans or latex beads moderately stimulate d keratinocyte growth.③Ker-atinocytes treated with intact C.albicans had significantly higher level of IL-1?in the supernatant but lower in the cell extract.Both TGF -?and bFGF increased either in the medi a or in the extract.Conclusion These results suggest that keratinocytes have non -specific phagocytic activ ity.C.albicans are able to adhere to ker-atinocytes and stimulate the release of various cytokines from keratinocytes,which may induce an inflammatory reaction and cell growth.

The effect of povidone iodine solution(PIS)on the metabolic activity of Candida albi- cans and the effect of PIS treated C.altn'cans to cultured human keratinocytes were studied by using the Alamar blue fluorescence assay.It was found that PIS inhibited the metabolic activity of Candida albi- cans with a dose-dependent manner,and that the LD_(50)of PIS for Candida albicans is 0.075%.After treating for 30 min by 2% PIS/PBS(-),Candida albicans almost lost the adherence and invasion to the cultured human keratinocytes,the wall of yeast cells became uneven and shrunken,the intracellular structures became obviously indistinct,and the yeasts could not develop to hypha form.Candida albi- cans,treated with 2% PIS in PBS(-)for 30 min or with 10% PIS Sabouraud's liquid medium for 33 hr,did not form colony when it was inoculated to Sabouraud's dextrose agar.This result suggests that the fungicidal effective of PIS is influenced by its solvent.

Objective To investigate the etiology of papular urticaria: is it caused only by arthropod-like insects-bite allergy, or by multiple factors such as food allergy, disturbance of gastrointestinal function and infection? Methods We searched, by computer and manually, the foreign and domestic literature related to the etiology of papular urticaria published since 1950s, and according to the methods of evidence-based medicine, systematically evaluated the evidence supporting either the insect-bite theory or the multiple factor theory. Results Twenty-nine articles ( 22 English and 7 Chinese ) supported the theory of hypersensitivity to bites from certain insects such as mosquitoes, gnats, fleas, mites, bedbugs etc. Two articles in Chinese mentioned the possibilities other than insect-bite, but the reliability was unconvincing, because the authors did not present the source of data or statistical methods used in the articles. The evidence from epidemiology, histopathology, laboratory and clinical researches all supported the insects-bite theory. No proven evidence was found supporting other aetiological hypotheses. Conclusion Our results suggest that papular urticaria is caused only by the allergy to stings or bites of arthropods, and other hypotheses still lack convincing evidence.

A 66-year-old man was admitted for a 7-day history of painful blisters and erosions in the upper lip.Real-time PCR with herpes simplex virus (HSV) type-specific primers showed that the blister fluid and crusts were positive for HSV-1,but negative for HSV-2.Observation of the blister wall with transmission electron microscopy (TEM) revealed 3 types of nucleocapsid in the karyoplast of epithelia cells,including the electron-translucent core,granular core and electron-dense core.Numerous matured viral particles with envelope were found in the cytoplasm,which were identified as HSV.The diagnosis was made as herpes simplex in the upper lip based on the above findings.PCR based molecular typing combined with observation of HSV particles via TEM may be an effective approach to the definite diagnosis of primary herpes simplex.

A strain of Prototheca species isolated from a case of meningitis was identified by routine morphologic and biochemical methods as well as amplification of the related genes, in which the 28S large-subunit (LSU) region of ribosomal RNA (rRNA) gene and intergenic space (ITS) were amplified with universal fungal primers. The small-subunit (SSU) rRNA gene was amplified with eukaryote-specific primers and Prototheca genus-specific primers. Then, compared the sequences with the ones posted on BLAST (www. ncbi. nlm. nih. gov/BLAST). The organism choice giving the closest match, up to 99%, was considered the most likely correct identification. It was found that this strain of fungus grew well at 25 ℃ or 37 ℃. Smooth,moist colonies with white color were observed on Sabouraud Dextrose Agar (SDA) and Potato Dextrose Agar (PDA). Microscopically, globular or ovoid cells, a number of round, ovoid shaped endospores could be observed. No hypha, ascus or blastic conidia was found upon cultivation on SDA. Based on the morphological characteristics, this isolate could be identified as Prototheca species. The identity with Prototheca wickerhamii was 2.9 % as demonstrated by the API 20C AUX system. Sequence analysis showed that the ITS gene was proved to be a complex structural region which was not suitable for the identification of Prototheca species, but the LSU and SSU rDNA regions showed 94% and 99.9% sequence identities with Prototheca zopfii var. hydrocarbonea (P. zopfii var. hydrocarbonea) respectively, indicating that the SSU rRNA gene sequence might be more reliable on than the LSU rRNA gene sequence for identification of Prototheca species.

Objective To combine the active poly lactic co-glycolic acid (PLGA) nanosperes with adriamycin in side (ADM-NP) on the surface of ultrasound micrbubbles in covalent bonding and to prepare a novel drug-loading ultrasound micrbubble,and observe the physicochemical property.To quantify drug encapsulation efficiency and drug-loading amounts and drug-release properties in vitro and the effect of tumor imaging.Methods ADM-NP were prepared with double emulsification method,The surface carboxyl of ADM-NP was activated with carbodiimide method.Amino ultrasound microbubbles (MB-NH2) were prepared with mechanical shaking.The carboxyl of ADM-NP and the MB-NH2 could take place condensation reaction under a certain condition.Light microscope was used to observe the shape and distribution of the novel micrbubble.High performance liquid chromatography (HPLC) was used to quantify drug encapsulation efficiency and drug loading amounts.This novel microbubbles were put in 2 dialysis bag to observe the releasing properties of ADM in vitro,and one of the bag was using low frequency ultrasound irradiation for 120 s.The effect of tumor imaging using this novel microbubble was observed.Results After 48 hours,a number of ADM-NP attacted to the MB-NH2 like a gar land.Determination of entrapment efficiency and drug loading of it by HPLC weare (86.11 ± 6.76)％ and (8.71 ± 0.46)％.The sustained release in vitro can last for more than 48 hours.More than 90％ of ADM encapsulated in the 2 groups was sustained released for 48 hours.And the release characteristics of the 2 groups in vitro was in accord with Higuchi equation,and no difference was observed in the 2 groups (P ＞0.05).The tumor showed typical enhancement pattems of quick wash-in and quick wash-out.Conclusions To combine the nanospheres to the surface of microbubbles with covalent bonding,it could prepare a novel efficient drug-loading microbubbles for a original technique of ultrasound-targeted microbubble destruction(UTMD).

Objective To investigate the distribution of Malassezia species in lesional and non-lesion-al sites of patients with pityriasis versicolor(PV),species-variation in different anatomic sites and in lesions with different pigmentation,and the relationship between various Malassezia species and severity and age of PV patients.Methods A total of629skin specimens taken by sterile adhesive tape from the lesions and non-lesional skin were inoculated on media containing rapeseed oil in113patients with PV.Isolated colonies were identified to species based on physiological and morphological characteristics.Results The isolation rates of Malassezia spp.were not significantly different from both lesions and corresponding non-lesional skin.Among non-lesional sites,the isolation rate was significantly higher in forehead and trunk than that in upper and lower extremities.Five species were identified out of565strains obtained from the patients,including M.sympodialis(44.78%),M.furfur(32.94%),M.globosa(11.68%),M.obtusa(5.84%)and M.restricta(4.76%).Two dif-ferent species were isolated simultaneously from27sites.There was no obvious difference in species distribu-tion patterns between lesions and non-lesional sites.M.restricta was isolated from forehead exclusively.Species-variation was closely linked to lesions with different pigmentation and the age of patients,not to the severity of disease.Conclusion There is neither statistical difference of Malassezia isolation rate and species distribution between lesions and non-lesional skin,nor correlation between disease severity and species-varia-tion.The anatomic sites,the diversity of pigmentation pattern and the age of patients seem to be associated with different Malassezia species.