The patient is an 11-year old boy, who was born with universe alopecia as well as dry and coarse skin. When he was 3 months old, photophobia was noticed, and since then, upper respiratory tract infection had occurred twice a month complicated by frequent diarrhea. He had short stature with slight conjunc- rival congestion, corneal vascularization, opacity, coarseness and poor vision. No abnormality was found in the teeth, sweating ability, or hearing. He had universal alopecia; his skin was dry and rough with generalized rhombus- or polygon-shaped scaly patches. Particularly thick brown scales were observed on the upper limbs. Moreover, there were spiny follicular papules on the abdomen and axillae, hyperkeratosis of palm and sole, and dystrophic nails. Hyperextensibility of proximal interphalangeal joints of the third, fourth and fifth fingers was noticed. He also suffered from mental retardation, the verbal intelligence quotient being 52, performance intelligence quotient lower than 40, full intelligence quotient lower than 40, but no abnormality was found in the heart, lung, liver or spleen. Histopathology of skin on the abdomen suggested a change characteristic of ichthyosis. Chromosome analysis revealed a karyotype of 46, XY. This is the first diagnosed case of ichthyosis follicularis with atrichia and photophobia syndrome in China.

Objective To investigate the DNA typing, observe relationship between DNA fingerprinting patterns and serotypes of Cryptococcus neoformans, and find a suitable genotyping standard for Cryptococcus neoformans. Methods Three primers, including CN 1(GTG) 5, CN 2(GACA) 4 and CN 3(GATA) 4, were used to distinguish variations among strains of C.neoformans. Results The distinguishable fingerprinting bands for serotype of C.neoformans were yielded by primer CN 2. Using this primer, of 24 clinical and environmental isolates of serotype A of C.neoformans and 8 standard strains investigated by PCR, 20 strains produced complete identical fingerprinting patterns, the other 4 strains had different fingerprinting patterns. 2 strains of serotype B and C yielded indistinguishable fingerprinting patterns. Conclusions ①The majority of strains of serotype A had similar and stable fingerprinting patterns. ②Some strains of serotype B and C had an indistinguishable fingerprinting patterns. ③The same serotype strains from different sources may produce different DNA fingerprinting patterns. ④The DNA fingerprinting is a rapid, simple and feasible method for identifying Cryptococcus neoformans.

This paper reports 20 patients with cryptococcal meningitis,which were misdiagnosed before anti-fungal therapy,and the majority of them was diagnosed as tuberculous meningitis or viral meningitis.These patients were treated either with amphotericin B or fluconazale alone,or in combina- tion with flucytosine respectively,of the 15 patients could be evaluated,10 were cured(66.7%),2 im- proved(13.3%),3 died(20%),1 relapsed(10%).The mortality rate among the under age group and the aged group was significantly higher than that of the adult group(71.4% vs 23%).It is the authors' experience that in the early stage intrathecal amphotericin B combines with intravenous fluoconazole and changes to oral fluconazole or itraconazole in the later stage may be a valuable approach for cryptococcal meningitis.

Objective To evaluate the efficiency of PCR-restriction fragment length polymorphism (PCR-RFLP)aiming at the structure gene g6341,versus PCR fingerprinting analysis in the genotyping of Cryptococcus neoformans MethodsEight reference strains and 68 clinical and environmental isolates of C.neoformans were genotyped by PCR-RFLP and PCR fingerprinfing.In PCR fingerprinting,the minisatel lite-specific core sequence of wild-type phage M13 was used as a single primer.The structure gene g6341 was selected for PCR-RFLP analysis by sequence alignments of multiple genes,a pair of pnmers were developed based on the conserved region of g6341 gene.PCR products were digested with the appropriate restriction endonucleases,and RFLP profiles were analyzed.Partial sequence analysis of g6341 gene was performed for different genotypes of C.Neoformans.Phylogenetic analysis was done to study the relatedness between these genotypes.Results As sequence homology analysis showed,g6341 gene was suitable for RFLP analysis.In the case of enotyping of 76 C. Neoformans strains,the results obtained from PCR-RFLP were consistent with those from PCR fingerprinting.Sequence analysis of g6341 gene revealed a homology of 84%-97%among the eight genotypes as well as a consistency of 99%-100%within a same genotype.In the phylogenetic tree,genotypes VNⅠ,VNⅡ,VNⅢand VNⅣ belonged to one cluster,and genotypes VGⅠ,VGⅡ,VGⅢ and VGⅣ to another cluster.Conclusions PCR-RFLP analysis aiming at the structure gene g6341 is a useful tool to genotype C.neoformans.Sequence analysis of g6341 gene can disclose the relatedness among different molecular types of C.neoformans.

Objective To identify the AD hybrid strains and its hybrid types within Cryptococcus neoformans.Methods Difierent hybrid types of AD strains were analyzed by PCR 0f STE20 and MF genes within MAT locus and CIA4 and GPal genes out of MAT locus.The PCR-RFLP analysis of g6341 gene was also performed.Results The mating types of 18 AD strains were precisely identified by PCR of STE20 gene,whereas those of H strain were not identified.CL44 gene was better than the GPal gene in PCR identification of the AD hybrids.In the RFLP analysis of g6341 gene,AD strains were grouped into 2 distinct RFLP patterns based on the mating type on serotype A allele.The mating types of AD strains were not identified by the molecular analyses based on the CL44,GPal and g6341 genes.Conclusion It is necessary to use multi-locus analyses of genes within and out of the MAT locus in precise identification of the AD strains and their hybrid types of Cryptococcus neoformans.

Objective To assess the subgenotypes of pathogenic Cryptococcus gattii isolates from China and to elucidate the epidemiological links between these domestic isolates and those from other parts of the world. Methods DNA was extracted from 9 clinical isolates of Ctyptococcus gattii from China. The partially variable regions of the three unlinked loci, namely IGS1, PLB1 and GEF1, were amplified and sequenced, and the bioinformation at these loci was obtained from GenBank for multi-locus sequences alignment and phylogenetic analysis. Results Of these 9 clinical isolates, 8 were genotype VG Ⅰ and mating type α with the same sequences at the tested regions as the reference strain WM276, which was a representative isolate of an independent subgenotype; 1 was of genotype VG Ⅱ and mating type α, which was the first report in China, with the tested sequences consistent with those of the referrence strain R272. Sequencing and phylogenetic analysis of GEF1 gene, which was located at mating type locus, successfully identified the genotypes and mating types of all the Cryptococcus gattii isolates involved here. Conclusions Multi-locus sequence analysis shows that causative Cryptococcus gattii isolates of genotype VG Ⅰ in China carry similar sequences at the tested loci in IGS1, PLB1 and GEF1 genes, to a widely distributed subgenotype in the world, and the sequences of the first VG Ⅱ genotype isolate from China resemble the less virulent subgenotype VG Ⅱ b found in Vancouver islands.

Objective To evaluate the role of Restriction fragment length polymorphism (RFLP) analysis in detection of the fragment of GEF1α/a gene which are both located at ct and a mating type loci in identification of species, varieties, genotypes and mating types of Cryptococcus neoformans species complex(Cryptococcus neoformans and Cryptococcus gattii). Methods The GEF1α/a gene was selected from 20 genes which both located at α and a mating type loci for RFLP analysis, according to the requirements of sequence similarities and primer design in PCR-RFLP analysis. Primer pair was designed from the conserved regions of GEF1α/a genes of distinct genotypes and mating types of reference strains to amplify a fragment of GEF1α/a gene from Cryptococcus neoformans and Cryptococcns gattii strains tested. Sequence alignment,restriction maps analysis, endonucleases selection and electrophoresis stimulation were conducted by using DNAMAN and Vector NTI software. EeoT14 Ⅰ and Hap Ⅱ endonucleases were selected for RFLP analysis of the GEF1α/a fragments amplified from 125 isolates of Cryptococcns neoformans and Cryptococcus gattii. Results An approximate 1 300 bp fragment was amplified from total 82 Cryptococcus neoformans and 43 Cryptoceccus gattii isolates. However, negative PCR results were found in the reference strains of Cryptococcus laurentii, Candida albicans, Candida tropicalis, Candida parapsilosis, Candida krnsei,Candida glabrata, Trichosporon asahii, Aspergillus fumigatns and Aspergillus flavus. RFLP analysis successfully identified the species, varieties, genotypes and mating types of total 125 isolates of Cryptococcus neoformans and Cryptococcns gattii tested in this study. Condusion PCR-RFLP analysis of the GEF1α/a fragment has the potential value in identification of species, varieties, genotypes and mating types of Cryptococeus neoformans species complex simultaneously and rapidly, and may be a useful tool in molecular epidemiological analysis.

Objective To investigate the molecular epidemiology of clinical cryptococcal isolates from China by analyzing the constituents and distributions of the varieties,genotypes and mating types (MAT)of them.Methods (1)PCR fingerprinting and PCR amplification were performed by using the minisatellite-specific core sequence of wild-type phage M13 as single primer.Genotypes of the 110 cryptococcal isolates from China were assigned by comparison with the reference strains of the 8 major molecular types loaded on gel.(2)Identification of the varieties and mating types was carried out by PCR using the specific primers of the varieties and mating types.Results Of the 110 clinical cryptococcal isolates,strains of Cryptococcus neoformans var.grubii with genetype VNⅠ and mating type MATα were the most representative ones(89.1%)followed by strains of C.neoformans var.gattii(8.2%)including isolates of genotype VG I,mating type MATα(7.3%)and genotype VGⅡ,mating type MATα(0.9%);AD hybrids with the genotype VNⅢ,mating type MAT-/α and genotype VN Ⅲ,mating type MATα/-(1.8%);and isolate of C.neoformans var.neoformans with the genotype VNⅣ and mating type MATa(0.9%).Conclusion Of the clinical isolates from China,all three varieties and AD hybrids are found.The vast majority(>99%) of strains possess the α allele in MAT locus and most of them are C.neoformans vat.grubii with the genotype VN I,which accord with the data of most studies of clinical molecular epidemiology in other geographic areas.However.no genotype of VNⅡ.VGⅢ and VGⅣ isolates are found in this study.

Objective To investigate the efficacy of autologous peripheral hematopoietic stem cell transplantation in the treatment of adult dermatomyositis. Methods A 21-year-old patient with dermatomyositis received autologous peripheral hematopoietic stem cell transplantation and was followed up for 6 years. Autologous peripheral hematopoietic stem cells were mobilized by recombinant human granulocyte colony stimulating factor (rhG-CSF) before the transplantation, and the conditioning regimens consisted of cyclophosphamide,methylprednisolone and cyclosporin. Rabbit anti-human T lymphocyte immunoglobulin began to be applied on day 3 after retransfer of stem cells. The improvement in symptoms, physical signs and biochemical indicators was observed, and hematopoietic restructuration and immunity resurrection were evaluated after the transplantation. Results After the transplantation, skin eruption greatly improved and gradually subsided. The muscle force of extremities restored from level Ⅳ before transplantation to level Ⅴ. The level of creatine kinase declined sharply after transplantation, but gradually returned to previous level. Leucocyte count began to decrease on the day of retransfer, and returned to the normal level on day 8. Immune function remained normal before and after the transplantation. Conclusion Autologous peripheral hematopoietic stem cell transplantation is an alternative treatment for severe and refractory dermatomyositis.

Objective To investigate the clinical features and analyze the molecular genetics of a pedigree of limb girdle muscular dystrophy (LGMD).Methods Pedigree analysis and clinical examination were performed in one four-generation family with LGMD.Electrophysiology and muscle biopsy were done in the affected members.With an informed consent, gene mutation, genome screening and linkage analysis were conducted in 26 members of this pedigree.Results Seven patients were identified.Pedigree analysis was consistent with autosomal dominant inheritance.Affected members had early presentation.Main features included proximal muscle weakness without dysarthria nor spasticity; electrophysiology and muscle biopsy revealed myopathic changes.LGMD1 A, 1B, 1C and facioscapulohumeral dystrophy genes were not detected by gene mutation analysis.Genome screening and linkage analysis did not reveal any linkage with the disease-causing gene and the reported loci of LGMD1D and LGMD1F genes.Conclusions The clinical manifestations of this LGMD family are highly heterogeneous, and the disease-causing gene of this family is not linked to any of the reported sites, suggesting this may be a new disease-causing locus, or a new genetic type of LGMD.