Detection of intracellular Helicobacter pylori in Candida. SPP from neonate oral swabs

SUMMARY BACKGROUND: There is evidence of detection of Helicobacter pylori (H. pylori) in the stool of newborns and in the yeast that colonizes the oral cavity of this age group. However, there is a lack of research to confirm it. This study proposes to determine the existence of the bacteria at an early age, specifically in newborns. OBJECTIVE: To identify intracellular H. pylori in oral yeasts and to detect antigens of the bacteria in newborn stools. METHODOLOGY: Cross-sectional and descriptive study. Samples were obtained from infants (oral swab and meconium). Identification of yeast species was performed using the following techniques: CHROMagar Candida, Germinal Tube Test and API Candida Identification System, then the yeasts were observed by light microscopy and fluorescence. Detection of H. pylori antigen in meconium and PCR were performed to amplify specific genes of the bacterium (rRNA16S, cagA, vacA s1a, vacA s1b, vacA s2, vacA m1, vacA m2 and dupA). RESULTS: Intracellular H. pylori was detected in yeast of the species Candida glabrata (C. glabrata) isolated from an oral swab of a newborn. CONCLUSION: The results of this study evidenced the existence of intracellular H. pylori in newborns.

RESUMO ANTECEDENTES: Há evidências de detecçâo de Helicobacter pylori (H. pylori) em fezes de recém-nascidos, como também dentro de leveduras que colonizam a cavidade oral dessa faixa etária. No entanto, faltam investigações que confirmem esses achados. OBJETIVO: Identificar H. pylori intracelular em leveduras de origem oral e detectar antígenos dessa bactéria em fezes neonatais. METODOLOGIA: Estudo transversal e descritivo. As amostras foram obtidas de bebês (zaragatoa oral e mecônio). As identificações das espécies de leveduras foram realizadas utilizando as seguintes técnicas: CHROMagar Candida, teste de tubo germinativo e sistema de identificação API Cândida. As leveduras foram observadas por microscopía óptica e fluorescência. Realizou-se a detecçâo de antígeno de H. pylori em mecônio e PCR para a amplificação de genes específicos desta bactéria (rRNA16S, cagA, vacA s1a, vacA s1b, vacA s2, vacA m1, vacA m2 e dupA). RESULTADOS: Foi detectado H. pylori intracelular em leveduras da espécie Candida glabrata (C. glabrata) isoladas a partir de zaragatoas oral de um recém-nascido. CONCLUSÃO: Os resultados deste estudo evidenciaram a existência interna de levedura de H. pylori em recém-nascidos.

Estudio clínico y micológico de otomicosis en diabéticos: una serie de 17 casos / Clinical and mycological study of otomycosis in diabetics: series of 17 cases

RESUMEN Introducción: La otomicosis en pacientes inmunosuprimidos esta caracterizada por ser bilateral y ser causada por candida. Pocos estudios comparan las características micológicas encontradas en la microscopía directa y el cultivo. Objetivo: Identificar las características clínicas y micológicas de la otomicosis en pacientes diabéticos. Material y método: Estudio transversal en centro hospitalario de segundo nivel. Criterios de inclusión: pacientes diabéticos con diagnóstico clínico de otomicosis. Intervención: la muestra se examinó directamente bajo el microscopio y se cultivó. Resultados: Se incluyeron 17 pacientes, 10 mujeres y 7 hombres con una edad media de 47,5 años. Los síntomas predominantes fueron hipoacusia en 91,4% (n =16), prurito en 82,4% (n =14), otorrea en 76,5% (n=13)y otalgia en 70,6% (n =12). Afección bilateral se encontró en 47,1% (n =8). Estudio directo al microscopio mostró levaduras en 94,1% (n =16) y 5,9% mostró aspergillus (n =1). Cándida fue el género más comúnmente encontrado en los cultivos y en el examen directo microscópico con 94,1% (n =16) y Candida albicans la especie más común con 88,2% (n =15). Conclusión: Candida albicans es el agente etiológico más común en pacientes diabéticos con otomicosis. Su presentación clínica más frecuente es hipoacusia, prurito y otorrea. El examen directo identificó adecuadamente a los géneros fúngicos.

ABSTRACT Introduction: Otomycosis in immunocompromised patients is characterize by its bilateral course and the predominant etiologic agent is Candida. Few studies compare the mycological features between microscopic direct exam and culture. Aim: To identify the clinical and mycological characteristics of otomycosis in diabetic patients. Material and method: Transversal study. Secondary care center. Inclusion criteria: diabetic patients with clinical diagnosis of otomycosis. Intervention: Direct examination under a microscope of the ear sample and culture. Results: We included 17 patients, 10 women, 7 men with a mean age of 47.5 years. Symptoms were hearing loss 94.1% (n = 16), pruritus 82.4% (n =14) otorrhoea 76.5% (n =13) and otalgia 70.6% (n =12). Bilateral involvement was found in 47.1% (n =8). Direct microscopic study found 94.1% of yeast (n =16) and 5.9% of Aspergillus (n =1). Candida was the most common fungal genus in culture and microscopic exam with 94.1% (n =16) of cases and Candida albicans was the most common species in 88.2% (n =15) cases. Conclusion: Candida albicans is the most common etiologic agent in diabetic patients with otomycosis. Main symptoms were hearing loss, itching and otorrhea. Direct exam correctly identified the fungal genus.

Candida glabrata among Candida spp. from environmental health practitioners of a Brazilian Hospital

Abstract The incidence of the species Candida albicans and non-albicans Candida was evaluated in a Brazilian Tertiary Hospital from the environment and health practitioners. In a 12-month period we had a total positivity of 19.65% of Candida spp. The most recurring non-albicans Candida species was C. glabrata (37.62%), generally considered a species of low virulence, but with a higher mortality rate than C. albicans. Subsequently, C. parapsilosis (25.74%) and C. tropicalis (16.86%) were the second and third most commonly isolated species. Considering the total samples collected from the emergency room and from the inpatient and the pediatric sector, 19.10% were positive for Candida spp., with the predominance of non-albicans Candida species (89.42%). The high percentage of positivity occurred in the hands (24.32%) and the lab coats (21.88%) of the health care assistants. No sample of C. albicans presented a profile of resistance to the drugs. All the non-albicans Candida species presented a decreased susceptibility to miconazole and itraconazole, but they were susceptible to nystatin. Most of the isolates were susceptible to fluconazole and amphotericin B. As expected, a high resistance rate was observed in C. glabrata and C. krusei, which are intrinsically less susceptible to this antifungal agent. The contamination of environmental surfaces by Candida spp. through hand touching may facilitate the occurrence of Candida infections predominantly in immunocompromised patients. In addition to that, the antifungal agents used should be carefully evaluated considering local epidemiologic trends in Candida spp. infections, so that therapeutic choices may be better guided.

Especies de levaduras aisladas en muestras de orina en un hospital regional de España / Isolated yeast species in urine samples in a Spanish regional hospital

La detección de candiduria en pacientes hospitalizados o inmunodeprimidos tiene una gran relevancia clínica. El objetivo de nuestro trabajo fue describir la frecuencia de aislamiento de diferentes especies significativas de levaduras en las muestras de orina procesadas en nuestro hospital en el periodo 2010-2013 y analizar su sensibilidad a los antifúngicos de uso habitual. La identificación de especies se realizó por siembra en un medio cromogénico, prueba de filamentación y sistemas automatizados (ASM Vitek y MALDI Biotyper), mientras que la sensibilidad a los antifúngicos se determinó con el sistema ASM Vitek. De los 632 aislamientos de levaduras obtenidos, 371 fueron Candida albicans y 261 especies de Candida no C. albicans. Las especies con mayor número de aislados resistentes fueron Candida glabrata y Candida krusei. Basados en nuestros resultados, consideramos que la identificación de la especie y el estudio de la sensibilidad a los antifúngicos deberían ser prácticas habituales por parte de los laboratorios cuando se aíslan especies diferentes a C. albicans

Candiduria detection in hospitalized or immunocompromised patients is of great clinical significance. The aim of our study was to describe the isolation frequency of significant species of yeasts in urine samples processed in our hospital during the period 2010- 2013, and to analyze their susceptibility to commonly used antifungal agents. Species identification was performed by seeding on a chromogenic medium, the filamentation test and automated systems (ASM Vitek and MALDI Biotyper), while susceptibility was determined using the ASM Vitek system. Of the 632 yeast isolates in urine, 371 were Candida albicans species and 261 non-C. albicans Candida spp. The species with the highest number of resistant isolates were Candida glabrata and Candida krusei. Based on the results obtained, we believe that species identification and the susceptibility study should be current practice in the laboratories when species other than C. albicans are isolated

Synergistic effects of tacrolimus and azole antifungal compounds in fluconazole-susceptible and fluconazole-resistant Candida glabrata isolates

In vitro interaction between tacrolimus (FK506) and four azoles (fluconazole, ketoconazole, itraconazole and voriconazole) against thirty clinical isolates of both fluconazole susceptible and -resistant Candida glabrata were evaluated by the checkerboard microdilution method. Synergistic, indifferent or antagonism interactions were found for combinations of the antifungal agents and FK506. A larger synergistic effect was observed for the combinations of FK506 with itraconazole and voriconazole (43%), followed by that of the combination with ketoconazole (37%), against fluconazole-susceptible isolates. For fluconazole-resistant C. glabrata, a higher synergistic effect was obtained from FK506 combined with ketoconazole (77%), itraconazole (73%), voriconazole (63%) and fluconazole (60%). The synergisms that we observed in vitro, notably against fluconazole-resistant C. glabrata isolates, are promising and warrant further analysis of their applications in experimental in vivo studies.

Occurrence of killer Candida glabrata clinical isolates

In this work we characterized the occurrence of killer activity in 64 Candida glabrata clinical isolates under different conditions. We found that only 6.25 % of the clinical isolates tested were positive for killer activity against a Saccharomyces cerevisiae W303 sensitive strain. Sensitivity of killer activity to different values of pH and temperatures was analyzed. We found that the killer activity presented by all isolates was resistant to every pH and temperature tested, although optimal activity was found at a range of pH values from 4 to 7 and at 37ºC. We did not observe extrachromosomal genetic elements associated with killer activity in any of the positive C. glabrata isolates. The killer effect was due to a decrease in viability and DNA fragmentation in sensitive yeast.

The activity of echinocandins, amphotericin B and voriconazole against fluconazole-susceptible and fluconazole-resistant Brazilian Candida glabrata isolates

The extensive use of azole antifungal agents has promoted the resistance of Candida spp to these drugs. Candida glabrata is a problematic yeast because it presents a high degree of primary or secondary resistance to fluconazole. In Brazil, C. glabrata has been less studied than other species. In this paper, we compared the activity of three major classes of antifungal agents (azoles, echinocandins and polyenes) against fluconazole-susceptible (FS) and fluconazole-resistant (FR) C. glabrata strains. Cross-resistance between fluconazole and voriconazole was remarkable. Among the antifungal agents, the echinocandins were the most effective against FS and FR C. glabrata and micafungin showed the lowest minimal inhibitory concentrations.

Candida spp: manual identification (reference method) and automated identification (Vitek system platform)

Yeasts are becoming a common cause of nosocomial fungal infections that affect immunocompromised patients. Such infections can evolve into sepsis, whose mortality rate is high. This study aimed to evaluate the viability of Candida species identification by the automated system Vitek-Biomerieux (Durham, USA). Ninety-eight medical charts referencing the Candida spp. samples available for the study were retrospectively analyzed. The system Vitek-Biomerieux with Candida identification card is recommended for laboratory routine use and presents 80.6% agreement with the reference method. By separate analysis of species, 13.5% of C. parapsilosis samples differed from the reference method, while the Vitek system wrongly identified them as C. tropicalis, C. lusitaneae or as Candida albicans. C. glabrata presented a discrepancy of only one sample (25%), and was identified by Vitek as C. parapsilosis. C. guilliermondii also differed in only one sample (33.3%), being identified as Candida spp. All C. albicans, C. tropicalis and C. lusitaneae samples were identified correctly.

Assessment of a real-time PCR method to detect human non-cryptococcal fungal meningitis

The signs and symptoms associated with fungal meningitis are similar to those seen with more common bacterial infections. In this study, we investigate whether Aspergillus or Candida DMA can be detected in cerebrospinal fluid [CSF] samples from patients suspected of fungal meningitis using real-time PCR assay. From April 2007 to November 2009, we evaluated CSF samples and sera from patients with risk factors for cerebral fungal meningitis in Nemazi Hospital, Shiraz University of Medical Sciences, Iran, by real-time PCR assay and routine mycological studies [direct microscopy examination and culture]. Two CSF and two serum samples from each patient were examined. CSF and serum samples from 38 patients [total: 152] suspected of fungal meningitis were examined. India ink staining and KOH smear were negative for all patients. C. albicans was isolated from two CSF samples. There were ten patients with positive real-time PCR results in their CSF samples: three patients had C. albicans, one with C. glabrata, four with Aspergillus species and two with both C. albicans and Aspergillus species DMA. Four patients had positive serum results for Aspergillus or Candida infections. Considering the findings, it seems that molecular examination can help in the diagnosis of fungal meningitis in patients with clinical and radiological presentations. Further studies should be conducted in other regions and settings to confirm these findings

Vinegar as an antimicrobialagent for control of Candida spp. in complete denture wearers

The use of denture is known to increase the carriage of Candida in healthy patients, and the proliferation of Candida albicans strains can be associated with denture-induced stomatitis. The aim of this study was to evaluate the use of vinegar as an antimicrobial agent for control of Candida spp. in complete upper denture wearers. Fifty-five patients were submitted to a detailed clinical interview and oral clinical examination, and were instructed to keep their dentures immersed in a 10%vinegar solution (pH less than 3) overnight for 45 days. Before and after the experimental period, saliva samples were collected for detection of Candida, counting of cfu/mL and identification of species by phenotypical tests (germ tube formation, chlamidoconidia production, and carbohydrate fermentation and assimilation). The results were analyzed using Spearman's correlation and Student's t-test (p£0.05). Candida yeasts were present in 87.3% of saliva samples before the treatment. A significant reduction was verified in CFU/mL counts of Candida after treatment. A positive correlation between Candida and denture stomatitis was verified, since the decrease of cfu/mL counts was correlated with a reduction in cases of denture stomatitis. Although it was not able to eliminate C. albicans, the immersion of the complete denture in 10% vinegar solution, during the night, reduced the amounts (cfu/mL) of Candida spp. in the saliva and the presence of denture stomatitis in the studied patients.

Candida Glabrata Esophagitis: new case reports and management / Esofagite por Candida Glabrata: novos relatos de casos e conduta terapêutica

Candida esophagitis (CE) is a common opportunistic infection in the immunocompromised host. C. glabrata is rarely cited as agent of CE and has been underestimated due to lack of proper identification. In this study, two cases of C. glabrata esophagitis in AIDS and chagasic patients are reported. Diagnosis of Candida species should be considered an important key for the ideal choice of antifungal therapy against this mycosis.

Esofagitepor Candida (CE) é uma infecção oportunista comum em hospedeiros imunocomprometidos. C. glabrata é raramente citada como agente de CE e tem sido subestimada devido à falta de uma identificação apropriada. Este estudo relata dois casos de esofagitepor C. glabrata em pacientes com AIDS e doença de Chagas. O diagnóstico das espécies de Candida deveria ser considerado uma importante chave para a escolha da terapia antifúngica ideal contra esta micose.

Comparación de diferentes métodos para la identificación de especies del género Candida / Comparison of different methods for species identification of genus Candida

Las diferentes especies del género Candida producen una variedad de enfermedades, desde infecciones mucocutáneas leves a formas diseminadas graves. Tradicionalmente, la taxonomía de las levaduras se ha llevado a cabo en base a estudios morfológicos y fisiológicos, pero éstos dependen de las condiciones de cultivo de las cepas, por lo que se han observado diversas dificultades. Por tal motivo, recientemente, se han probado técnicas de biología molecular. El objetivo de este trabajo es correlacionar los estudios taxonómicos de las especies correspondientes a las principales patógenas: C. albicans, C. krusei, C. parapsilosis, C. tropicalis y C. glabrata, realizados por técnicas fenotípicas tradicionales, métodos comerciales y por PCR fingerprinting. Al comparar las técnicas que identifican Candida albicans, agar harina de maíz y formación de tubos germinativos, estadísticamente se observa que no existen diferencias significativas entre ambos métodos (valor de la estadística X2 = 0,5 p = 0,4795). Comparando los métodos que discriminan especies de Candida: pruebas fisiológicas, CHROMagar, API20C y PCR fingerprinting se observó que no existen diferencias significativas en las proporciones de resultados que identifican cualquier Candida entre las pruebas fisiológicas, API20C y PCR fingerprinting. La proporción de resultados definitivos es mayor a la obtenida usando el método CHROMagar (p< 0,001).

Different species of genus Candida can cause a wide range of pathologies, since mucocutaneous trivial infections to disseminated serious forms. Traditionally, taxonomy of yeast has been performed taking into account morphologic and physiologic studies, but they depend on the culture conditions of strains, what cause certain difficulties. Thus, recently, molecular biology methods have been tried. The aim of this work is to correlate taxonomic studies of most important pathogenic species -C. albicans, C. krusei, C. parapsilosis, C. tropicalis and C. glabrata- all of them performed by phenotypic traditional methods, commercial ones, and by a molecular method, PCR fingerprinting. Comparing useful methods for C. albicans identification, corn flour agar and germinative tube formation, no statistical differences between them are observed (X2 =0.5, p=0.4795). By comparison between methods to discriminate different Candida species, physiological tests, CHROMagar, API 20C and PCR fingerprinting we observed no significative differences in proportion of accurate results, in test that can identify any Candida species, such as physiological assays, API 20C and PCR fingerprinting. The proportion of unequivocal results is greater than the obtained performing the CHROMagar culture method (p< 0.001).

Prevalencia de Candida albicans y Candida no albicans en diferentes muestras clínicas: Período 1999-2001 / Prevalence of Candida albicansand Candida no albicans inclinical samples during 1999-2001

Las levaduras implicadas en procesos patológicos son de indiscutible importancia debido al incremento experimentado por estas infecciones en las últimas décadas, a los cambios observados en las especies causales y al uso empírico de antifúngicos. En el Centro de Micología se estudiaron 1006 aislamientos provenientes de una amplia gama de muestras clínicas durante el periodo 1999-2001. Candida albicans con 40,3% resultó la especie de mayor frecuencia de aislamiento, pero las especies de Candida no albicans con 54,9% resultaron de mayor prevalencia y el 4,8% fueron otras levaduras. En los hemocultivos Candida parapsilosis con 34,9%, C. albicans con 30,2% y C. tropicalis con 25,6% resultaron las más recuperadas, mientras que C. glabrata se presentó con un 2,3%. En las secreciones mucosas C.albicans con 60%-80% fue la especie preponderante. Hemos detectado especies de Candida causantes de mediastinitis, lo que nos alerta sobre su importancia en estos procesos. Las infecciones del tracto urinario por levaduras se detectaron en mayor frecuencia en individuos hospitalizados, resultando C. albicans con 47,7% la especie más aislada, y dentro de Candida no albicans, C. glabrata con 24,8% y C. tropicalis con 20,0%. En las onixis candidiásicas C. parapsilosis con 37,7% desplazó a C.albicans con 22,0% de este lugar anatómico. Los estudios de sensiblidad al fluconazol de las especies de Candida nos permiten concluir que C.albicans es una especie sensible y que los mayores porcentajes de resistencia se observaron en C. glabrata (21,41%) y and C. krusei (69,23%).

The importance of epidemiological monitoring of yeasts involved in pathologic processes is unquestionable due to the increase of these infections over the last decade, the changes observed in species causing candidiasis, and empirical antifungal treatment. At the Mycology Center, 1006 isolates from a wide range of clinical samples were studied during 1999-2001. Candida albicans (40.3%) was the most isolated species, although, the Candida no albicans species with 54.9% showed the major prevalence. In blood cultures Candida parapsilosis (34.9%), C. albicans (30.2%) and C. tropicalis (25.6%) were recovered most frequently while C. glabrata represented only 2.3%. C. albicans with 60%-80% was the predominant specie in mucosal surface. We also detected Candida mediastinistis, which alert us over the importance at this location. Urinary tract infections caused by yeasts were more frequent in hospitalized patients, being C. albicans (47.7%), the most commonly isolated, followed by C. glabrata (24.8%) and C. tropicalis (20.0%). In the candidal onychomycoses, C. parapsilosis (37.7%) outplaced C. albicans (22.0%). Fluconazole susceptibility studies of Candida species allowed us to conclude that the majority of C. albicans islolates are susceptible, and that the highest resistance averages were observed in C. glabrata (21.41%) and C. krusei (69.23%).