Temporal similarity between Candida albicans genotypes in a Tunisian neonatal intensive care unit suggests several nosocomial cross-contamination episodes

The nosocomial transmission of Candida albicans in neonatal intensive care units (NICUs) is an increasing concern and understanding the route of this transmission is critical for adequate infection control measures. The aim of our study was to assess the likeliness of nosocomial acquisition of C. albicans in the NICU of Farhat Hached hospital in Sousse (Tunisia). We genotyped 82 isolates from 40 neonates and 7 isolates from 5 health care workers (HCWs) with onychomycosis, by using CDC3 microsatellite length polymorphism (MLP) and the high-resolution melting (HRM) analysis. Combined MLP and HRM CD3 analysis led to the delineation of 12 genotypes. Five temporal clustering caused by five genotypes occurred during the study period. Three of these genotypes were isolated in both neonates and HCWs. The first clustering included 28 isolates obtained between January 2003 and May 2004 from 16 neonates and 2 HCWs. The second clustering included three isolates collected in 2004 from three neonates and two HCWs. The third clustering included 11 isolates obtained from 6 neonates and 1 HCW in 2006. The two remaining clustering could not be associated with any HCW's contamination. These results argue for the nosocomial transmission of C. albicans in our NICU. The combined MLP and HRM analysis is a rapid first approach for tracking cross-contamination.     

Prevalence of specific and phylogenetically closely related genotypes in the population of Candida albicans associated with genital candidiasis in China

Genitourinary candidiasis, which is most frequently caused by Candida albicans, is a common problem worldwide. The pathogenesis of the infection, especially recurrence of the infection, remains to be elucidated. This study analyzed 199 independent Chinese C. albicans isolates using multilocus sequence typing (MLST) and microsatellite typing, with the focus on the isolates associated with vulvovaginal candidiasis (VVC) of Chinese women. MLST data of 221 vaginal isolates from other countries available from the consensus MLST database of C. albicans were retrieved for comparison. A total of 124 diploid sequence types (DSTs) were recognized from the Chinese C. albicans isolates, among which, 98 (79.0%) have not been reported in the MLST database of the species. The majority of the VVC (71.6%) and balanitis (92.3%) isolates from China were located in clade 1 of C. albicans; while only 40.6% of the vaginal isolates and 7.8% of the oral isolates from healthy volunteers were found in the same clade. Furthermore, 69.1% of the VVC and 84.5% of the balanitis isolates concentrated in a cluster of clade 1 with DST 79 as the primary founder. The isolates in this cluster possessed microsatellite genotypes CAI 30-45, CAI 32-46 and their close derivatives. Interestingly, a remarkable difference in genotype distribution patterns between Chinese and non-Chinese vaginal isolates of C. albicans was observed. Only 11.3% of the non-Chinese vaginal isolates compared were located in the cluster concentrated with Chinese VVC isolates. The results suggest significant association of specific and genetically similar genotypes with genital infections in China.     

Multicenter Brazilian study of oral Candida species isolated from AIDS patients

Oropharyngeal candidiasis continues to be considered the most common opportunistic disease in Aids patients. This study was designed to investigate species distribution, serotype and antifungal susceptibility profile among Candida spp. isolated from the oral cavity of Aids patients recruited from six Brazilian university centers. Oral swabs from 130 Aids patients were plated onto CHROMagar Candida medium and 142 isolates were recovered. Yeast isolates were identified by classical methods and serotyped using the Candida Check or target system-Iatron. Antifungal susceptibility testing was performed according to the NCCLS microbroth assay. C. albicans was the most frequently isolated species (91%), and 70% of the isolates belonged to serotype A. We detected 12 episodes of co-infection (9%), including co-infection with both serotypes of C. albicans. Non-albicans species were isolated from 12 episodes, 50% of them exhibited DDS or resistance to azoles. Otherwise, only 8 out 130 isolates of C. albicans exhibited DDS or resistance to azoles. Brazilian Aids patients are infected mainly by C. albicans serotype A, most of them susceptible to all antifungal drugs.     

Biotypes and randomly amplified polymorphic DNA (RAPD) profiles of subgingival Candida albicans isolates in HIV infection

A group of subgingival isolates of C. albicans recovered from Italian HIV-positive (HIV+) subjects were characterized both phenotypically and genotypically. Phenotyping of the isolates was carried out by a biotyping method based on the enzyme profiles, carbohydrate assimilation patterns and boric acid resistance of the yeasts. Genotyping was performed through randomly amplified polymorphic DNA (RAPD) analysis. Five biotypes were found among the 29 subgingival C. albicans strains examined. The predominant biotypes were A1R (55.17%), A1S (24.14%), and A2R (13.79%), while the biotypes A11R and A13R were represented by a single isolate each. RAPD profiles identified 15 genotypes among the 29 isolates. Almost every individual harboured his/her own specific isolate and in three out of the six subjects with multiple isolates (two to six each) more than one genotype (two to six) was found. The biotype distribution we found is consistent with previous reports on C. albicans isolates from other oral sources, whereas the resistance to boric acid was highly frequent in subgingival strains. RAPD analysis showed high genetic heterogeneity within subgingival isolates, also when isolates were phenotypically identical. These findings, obtained from HIV+ subjects living in Southern Italy, may be useful as baseline information on subgingival C. albicans colonization in the Mediterranean area.     

Examination of the genetic variability among biofilm-forming Candida albicans clinical isolates

Biofilms are microbial communities encased in a self-produced polymeric matrix and represent a common mode of microbial growth. Candida albicans is able to colonize the surface of catheters, prostheses, and epithelia, forming biofilms that are highly resistant to antimicrobial drugs. The objective of this study was the genotypic characterization of biofilm-forming C. albicans clinical isolates using RAPD (Random Amplified Polymorphic DNA). We have studied 25 clinical isolates of C. albicans from oral cavities, blood, skin, nail, stool, oesophagus biopsy and vaginal fluids from patients suffering from candidiasis. For each strain biofilm formation was analysed by measuring the ability to adhere to and grow on polystyrene plastic surfaces using XTT [2,3-bis(2-methoxi-4nitro-5sulfophenil)-2H tetrazolium-5carboxanilide] reduction assay. The similarity coefficients generated by RAPD using four different primers varied from 49 to 91%, indicating a high degree of genetic variability between the clinical isolates. The dendrogram clustered the isolates in four related groups, all groups included strains with very different abilities to form biofilms. The isolates with similar genotypes often showed very different biofilm formation abilities. Strains were grouped into clusters independently of their clinical sources. Our results suggested that a direct correlation does not exist between the biofilm-forming ability of natural populations of C. albicans and the genotype as determined by RAPD.     

Molecular subtyping of clinical isolates of <Emphasis Type="Italic">Candida albicans</Emphasis> and identification of <Emphasis Type="Italic">Candida dubliniensis</Emphasis>in Malaysia

The genotypes of 221 recent isolates of Candida albicans from various clinical specimens of 213 patients admitted to the University Malaya Medical Centre, Malaysia was determined based on the amplification of a transposable intron region in the 25 S rRNA gene. The analyses of 178 C. albicansisolated from nonsterile clinical specimens showed that they could be classified into three genotypes: genotype A (138 isolates), genotype B (38 isolates) and genotype C (2 isolates). The genotyping of 43 clinical isolates from sterile specimens showed that they belonged to genotype A (29 isolates), genotype B (10 isolates), genotype C (2 isolates) and genotype D (2 isolates). The overall distribution of C. albicans genotypes in sterile and nonsterile specimens appeared similar, with genotype A being the most predominant type. This study reported the identification of C. dubliniensis (genotype D) in 2 HIV-negative patients with systemic candidiasis, which were missed by the routine mycological procedure. The study demonstrated the genetic diversity of clinical isolates of C. albicans in Malaysia.     

新疆地区白念珠菌基因型分析及其体外药物敏感性研究

目的研究新疆地区汉族和维吾尔族患者来源的50株白念珠菌的基因型及其对两性霉素B、5-氟胞嘧啶、米卡芬净、伊曲康唑、氟康唑和咪康唑的体外敏感性。方法采用PCR法扩增白念珠菌rDNA 25S的Ⅰ类内含子包含区,根据扩增产物的大小判断基因型(A型为450 bp,B型为840 bp,C型为450 bp和840 bp)。采用CLSI M27-A液基微量稀释法测定50株白念珠菌对上述6种抗真菌药的体外敏感性。结果 50株菌分为3种基因型:A型30株,B型和C型各10株。所有菌株对两性霉素B、5-氟胞嘧啶、米卡芬净和咪康唑的MIC值较低,MIC范围依次为0.25～0.5μg/mL,0.125～0.5μg/mL,≤0.03μg/mL,0.25～8μg/mL;对伊曲康唑和氟康唑的MIC值较高,MIC范围分别为0.25～8μg/mL,0.5～64μg/mL。B型和C型对5-氟胞嘧啶的MIC值均为0.125μg/mL,对伊曲康唑和氟康唑的耐药率分别为84%、70%。不同族别来源的菌株基因型比较无显著差异(P>0.05),不同基因型菌株的抗真菌药物敏感性比较也无显著差异(P>0.05)。结论新疆地区白念珠菌分A,B,C三种基因型。汉...     

Identification of Candida albicans by polymerase chain reaction amplification of CaYST1 gene intron fragment

A single pair of primers, deduced from the intron nucleotide sequence of the Candida albicans CaYST1 gene, was used in PCR analysis performed with both genomic DNA and whole cells of clinical isolates of Candida species and other microorganisms. All the clinical C. albicans isolates generated the expected 310 bp amplicon; other Candida species as well as laboratory strains belonging to other fungal genera failed to amplify any DNA fragment, except for Candida pseudotropicalis (amplicon of 1200 bp), Kluyveromices marxianus (amplicon of 1250 bp) and Cryptococcus neoformans (several amplicons longer than 1200 bp). Unusual C. albicans isolates from Africa also yielded the expected 310 bp amplicon. These results indicate that genes containing intron sequences may be useful to design species-specific primers for identification of fungal strains by PCR. The sensitivity of the method was evaluated for C. albicans genomic DNA by using both various DNA concentrations (224 ng to 2.7 pg) and different cell amounts (10(7); to 5 cells). The results obtained may be useful in earlier detection of candidiasis.     

Characterization of Candida Species from Different Populations in Taiwan

The opportunistic Candida species existing as part of commensal microbiota in humans are usually the etiological agents causing infections. We investigated whether isolates collected from different age groups, hospital units, and sources have distinct characteristics. A total of 913 isolates comprising 395 Candida albicans, 230 Candida tropicalis, 202 Candida glabrata, 62 Candida parapsilosis, 13 Candida krusei, and 11 of other six species were analyzed. Urine was the most common source (41.2%), followed by sputum (16.3%), blood (15.2%), and others (27.3%). Candida albicans and C. parapsilosis were more prevalent in the working group [from 19 to 65 years], whereas C. tropicalis and C. glabrata were more prevalent in the elder one (≥ 66 years). We found that the age of patients and the source of isolates affect the distribution of species. On the other hand, the drug susceptibility of isolates was associated with fungal species and whether patients were hospitalized.     

Involvement of aspirin-sensitive oxylipins in vulvovaginal candidiasis

3(R)-Hydroxyoxylipins are produced via an aspirin-sensitive pathway in Candida albicans, an abundant pathogen in vulvovaginal candidiasis. In the present study, we have investigated the effect of aspirin on vaginal isolates of C. albicans from patients with recurrent candidiasis. Aspirin alone and with clotrimazole, a commonly used drug, strongly suppressed growth of C. albicans. 3(R)-Hydroxyoxylipins, which were selectively located in hyphae and other filamentous structures, but not in free blastospores, were almost totally suppressed by aspirin. Moreover, C. albicans stimulated prostaglandin E(2) (PGE(2)) production in HeLa cells. PGE(2) is a stimulus for germ tube formation in C. albicans. We conclude therefore that the administration of aspirin should be beneficial in the treatment of vulvovaginal candidiasis by dual ways: (i) by inhibition of 3(R)-hydroxyoxylipin formation, and (ii) by inhibition of PGE(2) formation in the infected host tissue.     

rDNA-based genotyping of clinical isolates of Candida albicans

The study presents an analysis of the restriction pattern ofrDNA fragments of 95 C. albicans isolates previously classified on the basis of the presence of the intron in rDNA into genotypes A (62 isolates), B (28), and C (5). Most isolates (61) with genotype A were classified as "subtype a" and one as "subtype d" (Karahan and Akar; 2005). No differences were observed in the restriction patterns of the tested genotype B isolates. Similarly, most genotype C strains (4/5) showed the same restriction pattern. The results indicate low subtyping variations of the analyzed isolates, which is in contrast to published data obtained from a Turkish collection of yeasts.     

Karyotyping of Candida albicans and Candida glabrata isolates from recurrent vaginal infections by pulsed-field gel electrophoresis

In the present study, 16 women with recurrent vulvovaginal candidiasis (RVVC) due to Candida albicans and Candida (Torulopsis) glabrata were followed for a period of 4 to 12 months, and 36 vaginal isolates were evaluated by pulsed-field gel electrophoresis (PFGE). Eleven women were infected by C. albicans and 5 by C. glabrata. Three electrophoretic karyotypes of C. albicans and 3 of C. glabrata were identified throughout the follow-up. All patients but one was infected with the same karyotype of C. albicans or C. glabrata during the follow-up period. Two different karyotypes of C. glabrata were identified in one patient in the course of 12 months. The results confirmed the diversity of the karyotypes of C. albicans and C. glabrata causing vulvovaginitis, and demonstrated the persistence of colonization with the same strain over different periods of time despite therapy (15/16 women).     

Point prevalence, microbiology and antifungal susceptibility patterns of oral Candida isolates colonizing or infecting Mexican HIV/AIDS patients and healthy persons

We have conducted a longitudinal study over a 3-year period to address the point prevalence, microbiological characteristics and antifungal susceptibility patterns of yeast isolates colonizing or infecting the oral cavities of 111 HIV-infected (51 adults, 60 children) and 201 non HIV-infected (109 adults, 92 children) Mexican persons. Regarding the epidemiology of oral candidiasis, Candida albicans was the most frequent species isolated. Seventy-one out of 85 isolates from colonized persons were C. albicans (83.5%), 27 isolates of them were from HIV-infected children and 44 from non HIV-infected patients. Sixty-two isolates belonged to serotype A which was the most prevalent serotype of C. albicans. Non-albicans species (Candida glabrata, Candida tropicalis and Candida parapsilosis, and Saccharomyces cerevisiae) were isolated from 16.5% of colonized patients and from 38.5% patients with candidiasis or Candida-related lesions. There were nine episodes of infection or colonization by at least 2 different yeast species. In the case of HIV/AIDS patients, it was determined that yeast carriage was not associated with the number of CD4+ cells or the viral load, but HAART reduced the prevalence of oral candidiasis. Overall, most patients harbored strains in vitro susceptible to fluconazole, however 10.8% of the yeasts were resistant to one or more azole antifungal agents and 29% were intermediate susceptible to them. On the contrary, 5-fluorocytosine was very active against all isolates tested, and amphotericin B was active against 97.9% of them.     

Molecular typing of Candida albicans isolates from patients and health care workers in a neonatal intensive care unit

Aims: The aim of this study was to investigate the genetic relatedness between Candida albicans isolates and to assess their nosocomial origin and the likeliness of cross‐transmission between health care workers (HCWs) and hospitalized neonates in a neonatal intensive care unit (NICU). Methods: We retrospectively analysed 82 isolates obtained from 40 neonates and seven isolates from onychomycosis of the fingers of five HCWs in a Tunisian NICU by using pulsed‐field gel electrophoresis (PFGE) and randomly amplified polymorphic DNA (RAPD) analysis with CA1 and CA2 as primers. Results: In RAPD analysis, the discriminatory power (DP) of CA1 and CA2 primers was 0·86 and 0·81, respectively. A higher DP was achieved by combining patterns generated by both primers (0·92), while PFGE karyotyping exhibited the lowest DP (0·62). The RAPD‐CA1/CA2 analysis revealed that 65·8% of isolates obtained from neonates derived from a limited number (6) of groups of genetically identical strains, that five temporal clusterings occurred during the study period and that three HCWs’ isolates and 11 isolates obtained from six neonates were identical. Conclusions: These findings argue for the nosocomial transmission of C. albicans in our NICU and for the transfer of strains from HCWs to patients. Significance and Impact of the Study: Identification of relatedness between Candida species obtained from neonates and health care workers by using molecular techniques with high discriminatory power is essential for setting up specific control measures in order to reduce the incidence of nosocomial candidiasis.     

Proteolytic activity of clinical Candida albicans isolates in relation to genotype and strain source

Proteolytic activity is regarded as one of the most important virulence factors of Candida albicans. Several authors recently demonstrated that some karyotypes and genotypes harbouring a group I self-splicing intron (CaLSU) located in the gene encoding the large rRNA subunit showed a high level of proteinase production. The aim of this study was to investigate the correlation between the level of proteinase production and the presence of the CaLSU intron in C. albicans isolates originating from the blood and respiratory tracts (sputum/pharyngeal swabs) of patients with and without oropharyngeal candidosis. The results revealed statistically significant differences in genotype distribution and the level of proteinase production between the C. albicans isolates obtained from blood and from the respiratory tract. Genotype A, without the intron, was prevalent in all groups of strains and its prevalence was higher among isolates from blood (75%) and from patients with candidosis (80%) compared with strains from colonisation (as opposed to infection) (57.8%). Isolates from blood produced significantly less proteinase than isolates from the respiratory tract (p<0.02), and this difference should be attributed to lower proteinase production of genotypes B and C from blood compared with genotypes B and C from the respiratory tract (p<0.01). The higher proteinase production of genotype B than of genotype A was found among respiratory tract isolates only. The presented data indicate that the association between proteinase production and the CaLSU intron depends on the strains' population. Further study is needed on well-defined groups of clinical isolates to elucidate whether the observed diversity in proteinase production plays a role in the selection of strains inducing bloodstream infections.     

Epidemiologic Typing of Methicillin-Resistant Staphylococcus aureus in Neonate Intensive Care Units Using Pulsed-Field Gel Electrophoresis

To elucidate the mode of dissemination of methicillin-resistant Staphylococcus aureus (MRSA) in neonate intensive care units (NICUs), a total of 223 isolates from 3 separate hospitals were investigated between 1994 and 1996 by a DNA fingerprinting technique with pulsed-field gel electrophoresis (PFGE). Exoprotein profiles of some strains were also examined using SDS-polyacrylamide gel-electrophoresis (SDS-PAGE) and the assessment of enzyme/toxin production such as coagulase, enterotoxin and TSST-1. Judging from the strain typing data from PFGE results and the epidemiological data, 2 different types of PFGE patterns (A and B) and their subtypes (A′, A″ and B′) were identified. The A type including A′ and A″ (comprising approximately 95% of the isolates) was markedly dominant. Only 5% of the isolates belonged to type B and subtype B′. On the other hand, MRSA isolated from adult patients admitted to the same hospital showed many different PFGE patterns. The results strongly suggested that some strain(s) with specific PFGE pattern(s) is prevalent in NICUs. Furthermore, isolates which expressed the same PFGE pattern did not always express the same SDS-PAGE pattern. There were some isolates with different abilities to produce coagulase, enterotoxin C and toxic-shock syndrome toxin (TSST)-1, and the abilities had no relation with a particular type of PFGE pattern. Therefore, a combination of PFGE analysis and biochemical analyses of coagulase, enterotoxin C and TSST-1 may provide us with more detailed information for the epidemiological study of MRSA in NICUs.     

Microbiological and immunological features of oral candidiasis

Candida albicans(C. albicans) is the major infectious agent of oral candidiasis, and both innate immunity and cell-mediated immune response participate in the control of the fungal infections. The aim of this study was to correlate the clinical forms of oral candidiasis with the number of colony forming units (CFU) of C. albicans in saliva and to characterize T cell response in patients with oral candidiasis. Participants included 75 subjects: 36 with lesions of candidiasis and 39 without lesions of oral candidiasis. A 2-ml sample of saliva was collected from all subjects for microbiological analysis. Cytokine levels were determined by ELISA in supernatants of peripheral blood mononuclear cells of 25 patients with oral candidiasis, after in vitro stimulation with C. albicans antigens. In 48% of patients, no association was observed with denture use. C. albicans was detected in the saliva of 91.7% of patients with oral candidiasis, and there was an association between the number of CFU and the presence of oral lesions. A type Th1 immune response was observed in supernatants of peripheral blood mononuclear cells stimulated with C. albicans antigens. In contrast, IL-5 and IL-10 levels were very low or undetectable. Together, this study shows an association between clinical forms of oral candidiasis and the number of colonies of C. albicans in saliva, and that a systemic immune response characterized by the production of TNF-alpha and IFN-gamma is observed in patients with oral candidiasis.     

Variable biotherapeutic effects of Lactobacillus acidophilus isolates on orogastric and systemic candidiasis in immunodeficient mice

Two commercially available isolates of Lactobacillus acidophilus (NCFM and LA-1) were compared for their capacities to protect immunodeficient bg/bg-nu/un and bg/bg-nu/+ mice from candidiasis. L. acidophilus NCFM prolonged survival of adult and neonatal bg/bg-nu/nu mice, inhibited disseminated candidiasis in both mouse strains, suppressed weight loss associated with Candida albicans infection in bg/bg-nu/nu females, but did not decrease the severity or the incidente of orogastric candidiasis in gnotobiotic mice. L. acidophilus LA-1 suppressed numbers of C. albicans in the alimentary tracts of bg/bg-nu/+ mice and reduced the severity of mucosal candidiasis in bg/bg-nu/nuand bg/bg-nu/+ mice; however, L. acidophilus LA-1 did not improve the survival of bg/bg-nu/nu mice after oral challenge (colonization) with C. albicansand it was associated with lethality in gnotobiotic adult female bg/bg-nu/nu mice. These results demonstrate that the two isolates of L. acidophilus differed in their capacity to protect immunodeficient mice from candidiasis.     

Comparison between efficacy of allicin and fluconazole against Candida albicans in vitro and in a systemic candidiasis mouse model

The efficacy of allicin compared with fluconazole in alleviating systemic Candida albicans infections was evaluated both in vitro and in vivo through a systemic candidiasis mouse model. Determination of in vitro minimum inhibitory concentrations (MICs) for different C. albicans isolates revealed that both allicin and fluconazole showed different MICs that ranged from 0.05 to 12.5 μg mL(-1) and 0.25 to 16 μg mL(-1) , respectively. A time-kill study showed a significant effect of allicin (P<0.01) against C. albicans, comparable to that of fluconazole. Scanning electron microscopy observation revealed that, similar to fluconazole, allicin produced structural destruction of C. albicans cell surface at low MIC and lysis or puncture at high MIC concentrations. Treatment of BALB/c mice systemically infected with C. albicans showed that although the allicin treatment (at 5 mg kg(-1) day(-1) ) was slightly less efficacious than fluconazole treatment in terms of the fungal load reduction and host survival time, it was still effective against C. albicans in terms of mean survival time, which increased from 8.4 to 15.8 days. These results demonstrate the efficacy of anticandidal effects of allicin both in vitro and in an animal model of candidiasis and affirm the potential of allicin as an adjuvant therapy to fluconazole.     

白念珠菌临床分离调查及基因分型研究

目的 了解白念珠菌临床分离情况,并探讨其药敏结果与基因分型的相关性.方法 回顾性分析本院2011年3～11月间临床分离白念珠菌分布及耐药性;随机选取232株,采用PCR方法扩增白念珠菌25S rDNA基因内含子区进行基因分型研究;采用ATB真菌药敏试剂条进行药敏分析;统计分析药敏结果与基因分型的相关性.结果 期间共检出酵母样真菌973例,占病原菌阳性样本数比率为15.7％ (973/6196);其中分离白念珠菌562株,占58％ (562/973),主要分布科室为呼吸科(39.1％)、老年科(13.2％)、ICU(7.7％)、神经内科(7.5％)、免疫科(6.0％)以及其他科室(26.5％);标本类型以下呼吸道为主(81.7％),其次为尿路(9.4％)、血液(1.8％)等.对氟胞嘧啶、两性霉素B、氟康唑、伊曲康唑及伏立康唑的耐药率分别为0.9％、0％、1.4％、1.6％和1.1％.随机选取的232株白念珠菌经PCR方法可分为3型:A型125株,B型96株,C型11株.各型在5种药物的耐药性上并无差异.结论 临床分离酵母样真菌以白念珠菌为主,感染部位以下呼吸道为主;临床分离株对5种抗真菌药物敏感度较高,主要基因型为A和B型,不同基因分型间药敏结果并无统计学差异.