Margarete Teresa Gottardo de Almeida

Detection of P. aeruginosa harboring bla CTX-M-2, bla GES-1 and bla GES-5, bla IMP-1 and bla SPM-1 causing infections in Brazilian tertiary-care hospital.

BackgroundNosocomial infections caused by Pseudomonas aeruginosa presenting resistance to beta-lactam drugs are one of the most challenging targets for antimicrobial therapy, leading to substantial increase in mortality rates in hospitals worldwide. In this context, P. aeruginosa harboring acquired mechanisms of resistance, such as production of metallo-beta-lactamase (MBLs) and extended-spectrum beta-lactamases (ESBLs) have the highest clinical impact. Hence, this study was designed to investigate the presence of genes codifying for MBLs and ESBLs among carbapenem resistant P. aeruginosa isolated in a Brazilian 720-bed teaching tertiary care hospital.MethodsFifty-six carbapenem-resistant P. aeruginosa strains were evaluated for the presence of MBL and ESBL genes. Strains presenting MBL and/or ESBL genes were submitted to pulsed-field gel electrophoresis for genetic similarity evaluation.ResultsDespite the carbapenem resistance, genes for MBLs (blaSPM-1 or blaIMP-1) were detected in only 26.7% of isolates. Genes encoding ESBLs were detected in 23.2% of isolates. The blaCTX-M-2 was the most prevalent ESBL gene (19.6%), followed by blaGES-1 and blaGES-5 detected in one isolate each. In all isolates presenting MBL phenotype by double-disc synergy test (DDST), the blaSPM-1 or blaIMP-1 genes were detected. In addition, blaIMP-1 was also detected in three isolates which did not display any MBL phenotype. These isolates also presented the blaCTX-M-2 gene. The co-existence of blaCTX-M-2 with blaIMP-1 is presently reported for the first time, as like as co-existence of blaGES-1 with blaIMP-1.ConclusionsIn this study MBLs production was not the major mechanism of resistance to carbapenems, suggesting the occurrence of multidrug efflux pumps, reduction in porin channels and production of other beta-lactamases. The detection of blaCTX-M-2,blaGES-1 and blaGES-5 reflects the recent emergence of ESBLs among antimicrobial resistant P. aeruginosa and the extraordinary ability presented by this pathogen to acquire multiple resistance mechanisms. These findings raise the concern about the future of antimicrobial therapy and the capability of clinical laboratories to detect resistant strains, since simultaneous production of MBLs and ESBLs is known to promote further complexity in phenotypic detection. Occurrence of intra-hospital clonal dissemination enhances the necessity of better observance of infection control practices.

High Prevalence of bla(CTX-M) Extended Spectrum Beta-Lactamase Genes in Klebsiella pneumoniae Isolates from a Tertiary Care Hospital: First report of bla(SHV-12), bla(SHV-31), bla(SHV-38), and bla(CTX-M-15) in Brazil

The aim of this study was to investigate the presence and prevalence of bla(TEM), bla(SHV), and bla(CTX-M) and bla(GES)-like genes, responsible for extended spectrum beta-lactamases (ESBLs) production in clinical isolates of Klebsiella pneumoniae collected from a Brazilian tertiary care hospital. Sixty-five ESBL producing K. pneumoniae isolates, collected between 2005 and 2007, were screened by polymerase chain reaction (PCR). Identification of bla genes was achieved by sequencing. Genotyping of ESBL producing K. pneumoniae was performed by the enterobacterial repetitive intergenic consensus-PCR with cluster analysis by the Dice coefficient. The presence of genes encoding ESBLs was confirmed in 59/65 (90.8%) isolates, comprising 20 bla(CTX-M-2), 14 bla(CTX-M-59), 12 bla(CTX-M-15), 9 bla(SHV-12), 1 bla(SHV-2), 1 bla(SHV-2a), 1 bla(SHV-5), and 1 bla(SHV-31) genes. The ESBL genes bla(SHV-12), bla(SHV-31), and bla(CTX-M-15), and the chromosome-encoded SHV-type beta-lactamase capable of hydrolyzing imipenem were detected in Brazil for the first time. The analysis of the enterobacterial repetitive intergenic consensus-PCR band patterns revealed a high rate of multiclonal bla(CTX-M) carrying K. pneumoniae isolates (70.8%), suggesting that dissemination of encoding plasmids is likely to be the major cause of the high prevalence of these genes among the K. pneumoniae isolates considered in this study.

Caracterização fenotípica de leveduras isoladas da mucosa vaginal em mulheres adultas

PURPOSE to characterize, phenotypically, yeasts isolated from the vaginal content of 223 symptomatic (S) and asymptomatic (A) adult women with vulvovaginitis, and to determine the clinical indicators which may lead to the appearance of signs and symptoms related to the mucosa involvement by this pathology. METHODS a questionnaire with open and closed questions on epidemiological clinical data was applied initially. Then, mycological diagnosis with sowing in Chrom Agar Candida was done, followed by micro-morphological and biochemical identification. Specific methods for the detection of the virulence factors, proteinase and phospholipase were employed. Statistical analysis was performed through chi2 and Pearsons chi2 tests. RESULTS the most prevalent species found was Candida albicans (87%, S and 67%, A) followed by Candida glabrata (4%, S e 17% A). The number of women reporting the use of contraceptives was higher among the symptomatic, 77%. In the two groups studied, about 87% of the women presented regular menstrual cycles and 57% were married with ages between 30 to 40 years old. Concerning the sexual practices, there has been concomitance among anal, oral and vaginal habits from the patients. Only Candida albicans produced the virulence factor phospholipase in 37.5% of them. Proteinase has been detected in Candida albicans, Candida glabrata and Candida parapsilosis. This latter virulence factor was mainly associated to isolates from symptomatic patients. CONCLUSIONS it is a fact that the vaginal mucosa can be colonized and infected by yeasts, with several Candida species present. Nevertheless, Candida albicans is the most prevalent in the vaginal mucosa of adult women. It is evident the emergence of non-albicans Candida species, some of them with intrinsic resistance to azolics, such as Candida glabrata, Candida parapsilosis, Candida tropicalis, and Candida guillermondii, which can be explained by the inadequate use of medicines and empirical treatment.

Onicomicose: estudo clínico, epidemiológico e micológico no município de São José do Rio Preto

This was a clinical-epidemiological and mycological study on 184 patients at the university hospital: 200 samples, 142 positive samples, 98 yeasts and 68 filamentous fungi. Candida parapsilosis (47%) and Trichophyton rubrum (38%) were prevalent. They were 100% sensitive to ketoconazole and 99% sensitive to amphotericin B. The highest prevalences were among women (80%), adults (62%) and toes (84%).

MLST-Based Population Genetic Analysis in a Global Context Reveals Clonality amongst Cryptococcus neoformans var. grubii VNI Isolates from HIV Patients in Southeastern Brazil

Cryptococcosis is an important fungal infection in immunocompromised individuals, especially those infected with HIV. In Brazil, despite the free availability of antiretroviral therapy (ART) in the public health system, the mortality rate due to Cryptococcus neoformans meningitis is still high. To obtain a more detailed picture of the population genetic structure of this species in southeast Brazil, we studied 108 clinical isolates from 101 patients and 35 environmental isolates. Among the patients, 59% had a fatal outcome mainly in HIV-positive male patients. All the isolates were found to be C. neoformans var. grubii major molecular type VNI and mating type locus alpha. Twelve were identified as diploid by flow cytometry, being homozygous (AαAα) for the mating type and by PCR screening of the STE20, GPA1, and PAK1 genes. Using the ISHAM consensus multilocus sequence typing (MLST) scheme, 13 sequence types (ST) were identified, with one being newly described. ST93 was identified from 81 (75%) of the clinical isolates, while ST77 and ST93 were identified from 19 (54%) and 10 (29%) environmental isolates, respectively. The southeastern Brazilian isolates had an overwhelming clonal population structure. When compared with populations from different continents based on data extracted from the ISHAM-MLST database (mlst.mycologylab.org) they showed less genetic variability. Two main clusters within C. neoformans var. grubii VNI were identified that diverged from VNB around 0.58 to 4.8 million years ago.

Enteropathogens associated with acute diarrheal disease in children

OBJECTIVE: Aiming a better understanding of the distribution of enteropathogens in Brazil, we have investigated the prevalence of enteropathogens in acute infantile diarrhea in children of low economic level from São José do Rio Preto, SP.METHODS: A prospective study with 196 children under age five, all with diarrhea, and 33 control subjects was conducted at Hospital de Base de São José do Rio Preto during 1995. This Hospital is associated to a Medicine School and offers free assistance to patients of low economic level who live in the area. RESULTS: Enteropathogens were identified in 48% of the cases and 27% of the controls. Shigella species were isolated in 21% of the cases, EPEC in 10.7%, Salmonella species in 5.6%. The other enteropathogens studied represented 12 % of the isolate pathogens from case children. In the controls 18.2% of atypical EPEC were isolated. Isolation of Shigella species increased with increasing age of cases and peaked in spring, whereas EPEC was common in early infancy and peaked in spring and winter.CONCLUSIONS: Among São José do Rio Preto children, Shigella species and diarrheiogenic E. coli strains (EPEC, ETEC, EAggEC, EIEC) were isolated throughout the year as a cause of diarrhea bringing children to a medical attention. Most of the pathogens were isolated in spring (mainly Shigella and Salmonella), so the temperature and humidity of the environment must be very important.

Identification of fungal diseases at necropsy

The purpose of the Death Verification Service is to elucidate the causes of deaths that occur without medical assistance and of ill-defined deaths. In recent decades, the epidemiological reality of fungal infections has changed due to the rise in opportunistic infections chiefly in immunocompromised patients. A study of fungal diseases in autopsies performed in the Death Verification Service of the Medicine School in São José do Rio Preto between January 2000 and December 2009 was made. Sixty-seven cases of fungal disease, most involving men (70%), were found in 4824 autopsies. Cryptococcosis was the most prevalent (45%), followed by paracoccidioidomycosis, candidiasis, histoplasmosis, aspergillosis and mucormycosis. Associations between AIDS (n=14) and fungal diseases were identified for cryptococcosis (36%), candidiasis (28.5%) and histoplasmosis (28.5%). Pneumonia, AIDS and fungal diseases were evident in 26% of the cases, with the most prevalent etiologies being Cryptococcus neoformans (55.5%) and Histoplasma capsulatum (22%). Pneumonia alone occurred in 43% of cases, with cryptococcosis (53%) and paracoccidioidomycosis (33%) being the main infectious agents. Diabetes mellitus was associated with candidiasis in two cases and aspergillosis in one. One case of renal transplantation linked to paracoccidioidomycosis and one case of bone marrow aplasia with mucormycosis were reported. Despite the reduction in the number of autopsies over recent decades, these findings suggest that this procedure is useful to provide additional data on the etiology, underlying disease and specific risk factors, essential for quality control and to improve treatment protocols.

Trends in bacterial resistance in a tertiary university hospital over one decade

The objective of this study was to investigate bacterial resistance trends, infection sites and the relationship between resistance and admittance to the intensive care unit (ICU). A total of 53,316 bacteria identified between 1999 and 2008 were evaluated. Multidrug resistance was characterized when gram-negative bacilli (GNB) presented resistance to two or more classes of antibiotics. Gram-positive cocci (CPC) were assessed for resistance to penicillin, oxacillin and vancomycin. GNB were the most common (66.1%) isolate. There was a 3.7-fold overall increase in multidrug resistant GNB over the study period; Acinetobacter baumanii and Staphylococcus aureus were the most prevalent. Highest increases were recorded for Klebsiella pneumoniae (14.6-fold) and enterococci (73-fold). The resistance rates for GNB and GPC were 36% and 51.7%, respectively. Most multidrug resistant GNB and GPC were recovered from ICU patients (p-value<0.001). Vancomycin-resistant enterococci were isolated during this decade with an increase of 18.7% by 2008. These data confirm the worldwide trend in multidrug bacterial resistance.

First report of Tritirachium oryzae infection of human scalp.

The natural habitat of Tritirachium oryzae is soil and decaying plant material. It is also an insect pathogen. As a human pathogen, it has been reported as a cause of corneal ulcers and otomycosis. The case of a 4-year-old infant is reported with Tritirachium oryzae infection of the scalp. Diagnosis was established by direct mycological study and culture that showed Tritirachium oryzae as the only agent in a pure culture. The topical treatment involved an antifungal medication to a complete cure. We report the first case of scalp dermatomycosis due to Tritirachium oryzae infection, illustrating a novel clinical manifestation.

Essential Oil of Cymbopogon nardus (L.) Rendle: A Strategy to Combat Fungal Infections Caused by Candida Species

Background: The incidence of fungal infections, especially those caused by Candida yeasts, has increased over the last two decades. However, the indicated therapy for fungal control has limitations. Hence, medicinal plants have emerged as an alternative in the search for new antifungal agents as they present compounds, such as essential oils, with important biological effects. Published data demonstrate important pharmacological properties of the essential oil of Cymbopogon nardus (L.) Rendle; these include anti-tumor, anti-nociceptive, and antibacterial activities, and so an investigation of this compound against pathogenic fungi is interesting. Objective: The aim of this study was to evaluate the chemical composition and biological potential of essential oil (EO) obtained from the leaves of C. nardus focusing on its antifungal profile against Candida species. Methods: The EO was obtained by hydrodistillation and analyzed by gas chromatography-mass spectrometry (GC-MS). Testing of the antifungal potential against standard and clinical strains was performed by determining the minimal inhibitory concentration (MIC), time-kill, inhibition of Candida albicans hyphae growth, and inhibition of mature biofilms. Additionally, the cytotoxicity was investigated by the IC50 against HepG-2 (hepatic) and MRC-5 (fibroblast) cell lines. Results: According to the chemical analysis, the main compounds of the EO were the oxygen-containing monoterpenes: citronellal, geranial, geraniol, citronellol, and neral. The results showed important antifungal potential for all strains tested with MIC values ranging from 250 to 1000 μg/mL, except for two clinical isolates of C. tropicalis (MIC > 1000 μg/mL). The time-kill assay showed that the EO inhibited the growth of the yeast and inhibited hyphal formation of C. albicans strains at concentrations ranging from 15.8 to 1000 μg/mL. Inhibition of mature biofilms of strains of C. albicans, C. krusei and C. parapsilosis occurred at a concentration of 10× MIC. The values of the IC50 for the EO were 96.6 μg/mL (HepG-2) and 33.1 μg/mL (MRC-5). Conclusion: As a major virulence mechanism is attributed to these types of infections, the EO is a promising compound to inhibit Candida species, especially considering its action against biofilm.