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Glomerella leaf spot (GLS) is one of the most important diseases of apple, affecting a wide range of economically important cultivars, particularly Golden Delicious and its descendants. Caused mainly by species of the Colletotrichum gloeosporioides species complex (CGSC), C. fructicola has been described as the most prevalent and aggressive species associated with GLS and apple bitter rot (ABR) in Brazil and Uruguay. Recently, new CGSC species, closely related to C. fructicola, have been identified causing ABR. To verify the accuracy of species identification within the CGSC, we aimed to reevaluate the identity of representative GLS-causing isolates from Brazilian and Uruguayan populations, previously identified as C. fructicola. Multilocus phylogenetic analysis based on APN2, ApMAT, CAL, GAPDH, GS, ITS, and TUB2 allocated these isolates in a monophyletic clade with C. chrysophilum. This species was first described as the causal agent of anthracnose in banana fruits in Brazil, and recent reports indicate its association with ABR in the United States. This is the first report of C. chrysophilum causing GLS disease on apple worldwide.
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Colletotrichum , Malus , Brasil , Besouros , Colletotrichum/genética , Phyllachorales , Filogenia , Doenças das Plantas , UruguaiRESUMO
We present the first worldwide study on the apple (Malus × domestica) fruit microbiome that examines questions regarding the composition and the assembly of microbial communities on and in apple fruit. Results revealed that the composition and structure of the fungal and bacterial communities associated with apple fruit vary and are highly dependent on geographical location. The study also confirmed that the spatial variation in the fungal and bacterial composition of different fruit tissues exists at a global level. Fungal diversity varied significantly in fruit harvested in different geographical locations and suggests a potential link between location and the type and rate of postharvest diseases that develop in each country. The global core microbiome of apple fruit was represented by several beneficial microbial taxa and accounted for a large fraction of the fruit microbial community. The study provides foundational information about the apple fruit microbiome that can be utilized for the development of novel approaches for the management of fruit quality and safety, as well as for reducing losses due to the establishment and proliferation of postharvest pathogens. It also lays the groundwork for studying the complex microbial interactions that occur on apple fruit surfaces.
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Malus , Microbiota , Bactérias/genética , Frutas/microbiologia , Fungos/genética , Malus/microbiologiaRESUMO
Peach (Prunus persica L.) is an economically important deciduous fruit crop in Uruguay. Anthracnose caused by species of the genus Colletotrichum is one of the major diseases in peach production, originating significant yield losses in United States (Hu et al. 2015), China (Du et al. 2017), Korea (Lee et al. 2018) and Brazil (Moreira et al. 2020). In February 2017, mature peach fruits cv. Pavia Canario with symptoms resembling anthracnose disease were collected from a commercial orchard located in Rincon del Colorado, Canelones, in the Southern region of Uruguay. Symptoms on peach fruit surface were characterized as circular, sunken, brown to dark-brown lesions ranging from 1 to 5 cm in diameter. Lesions were firm to touch with wrinkled concentric rings. All lesions progressed to the fruit core in a V-shaped pattern. The centers of the lesions were covered by orange conidial masses. Monosporic isolates obtained from the advancing margin of anthracnose lesions were grown on PDA at 25ºC and 12h photoperiod under fluorescent light. The representative isolates DzC1, DzC2 and DzC6 were morphologically and molecularly characterized. Upper surface of colonies varied from white or pale-gray to gray and on the reverse dark-gray with white to pale-gray margins. Conidia were cylindrical, with both ends predominantly rounded or one slightly acute, hyaline and aseptate. The length and width of conidia ranged from 9.5 to 18.9 µm (x Ì =14.1) and from 3.8 to 5.8 µm (x Ì =4.6), respectively. The ACT, ßTUB2, GAPDH, APN2, APN2/MAT-IGS, and GAP2-IGS gene regions were amplified and sequenced with primers ACT-512F/ACT-783R (Carbone and Kohn, 1999), BT2Fd/BT4R (Woudenberg et al. 2009), GDF1/GDR1 (Guerber et al. 2003), CgDLR1/ColDLF3, CgDLF6/CgMAT1F2 (Rojas et al. 2010) and GAP1041/GAP-IGS2044 (Vieira et al. 2017) respectively and deposited in the GenBank database (MZ097888 to MZ097905). Multilocus phylogenetic analysis revealed that Uruguayan isolates clustered in a separate and well supported clade with sequences of the ex-type (isolate ICMP 18578) and other C. siamense strains (isolates Coll6, 1092, LF139 and CMM 4248). To confirm pathogenicity, mature and apparently healthy peach fruit cv. Pavia Canario were inoculated with the three representative isolates of C. siamense (six fruit per isolate). Fruit were surface disinfested with 70% ethanol and wounded with a sterile needle at two equidistant points (1 mm diameter x 1 mm deep). Then, fruit were inoculated with 5 µl of a spore suspension (1×106 conidia mL-1) in four inoculation points per fruit (two wounded and two unwounded). Six fruit mock-inoculated with 5 µl sterile water were used as controls. Inoculated fruit were placed in moist chamber and incubated at 25°C during 10 days. Anthracnose lesions appeared at 2 and 4 days after inoculation in wounded and unwounded points, respectively. After 7 days, disease incidence was 100% and 67% for wounded and unwounded fruit, respectively. The control treatment remained symptomless. The pathogens were re-isolated from all lesions and re-identified as C. siamense. C. siamense was previously reported in South Carolina causing anthracnose on peach (Hu et al. 2015). To our knowledge, this is the first report of anthracnose disease on peach caused by C. siamense in Uruguay. Effective management strategies should be implemented to control anthracnose and prevent the spread of this disease to other commercial peach orchards.
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Apple (Malus domestica Borkh.) is an important fruit crop in Río Negro, Argentina. In recent years, the frequency of canker and dieback symptoms have increased affecting different apple cultivars. In September 2014, a higher occurrence of cankers (29%) and dieback (9%) was observed in a commercial orchard of 10-year-old apple trees (n=210) cv 'Ital Red' in General Roca, Río Negro, Argentina (39°2'36.73"S - 67°32'44.55"W). Symptoms initially appeared as necrotic bark lesions on tree trunks and branches often associated with pruning wounds. Superficially, papyrus detachment of the bark and cracked bark were observed on the affected area. When the bark was removed, the diseased wood showed dark brown color. Cross sections of diseased branches revealed necrotic lesions that progress to branch death. Samples were collected from different symptomatic trees (n=30) and were superficially disinfected with 70% ethanol. Internal tissues (0.5 cm2) were excised from the advance margin of the necrotic lesions, plated on potato dextrose agar (PDA), and incubated at 22°C. Pycnidia were induced on sterilised pine needles overlaid on 2% water agar under near-UV light. Optimum temperature of culture growth on PDA was studied. According to their morpho-cultural characteristics, three different morphotypes were identified. The first, showed optimum growth at 30°C, had moderately dense white aerial mycelia and turned dark gray after 7 d. Conidia were ovoid, mostly aseptate, 20.8-25.6 × 8-11.4 µm (n=50) and hyaline to brown. The second, exhibited optimum growth from 25 to 30°C, was white to gray, with sparse to moderate aerial mycelium that turned dark olive green. Conidia were ovoid, 1-septate, 17.6-22.4 × 8.1-11.2 µm (n=50) and brown. Finally, the third showed optimum growth at 25°C, mycelium was grey to dark olive green. Conidia were oblong to ovoid with both ends rounded, aseptate and 1-septate at maturity, 20.8-24 × 11.2-14.4 µm (n=50), hyaline turned brown. Genomic DNA was extracted from one representative isolate of each morphotype and the ITS and EF1-α loci were amplified with the primer sets ITS1/ITS4 (White et al., 1990) and EF1-728F/EF1-986R (Carbone and Kohn, 1999), respectively. The nucleotide sequences indicated ≥99% identity to D. seriata (CBS 114796 and CBS 112555), D. mutila (CBS 302.36 and CBS 112553), and D. juglandis (CBS 188.87), reclassified as Dothiorella omnivora (Linaldeddu et al., 2016), for both DNA regions. The sequences were deposited in the GenBank database (MW596418, MW598375; MH665432, MK955889; MH665413, MK937229). To confirm pathogenicity, healthy 1-year-old twigs of adult apple trees were pruned and wounds of attached twigs immediately inoculated with 20 µL of conidial suspension (103 conidia.mL-1, n=9 per isolate) or sterile distilled water (control, n=9), and wrapped with Parafilm. The experimental design was randomized, and the experiment was repeated once. After 90 d, the area of lesion on all twigs inoculated was determined. D. mutila and Do. omnivora produced mean canker areas (65 and 73 mm2, respectively) significantly larger (p < 0.005) than D. seriata (48 mm2). No lesion occurred in the negative controls. Fulfilling Koch's postulates, fungi were reisolated from all inoculated twigs and no fungus was recovered from controls. To our knowledge, this is the first report of D. seriata, D. mutila, and Do. omnivora associated with apple canker and dieback in Argentina, which shows the need of study the role of these fungi in orchard health.
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Colletotrichum fructicola is the main species causing apple bitter rot (ABR) and Glomerella leaf spot (GLS) in southern Brazil, and ABR in Uruguay where GLS remains unnoticed. Thus, this work aimed to determine the genetic structure of C. fructicola isolates of both the countries. A total of 28 out of 31 Brazilian isolates (90.3%) caused typical symptoms of GLS, while only 6 of 25 Uruguayan isolates (24.0%) originating from fruits were able to infect leaves, but causing atypical symptoms. Both populations showed similar levels of Nei's gene diversity (h = 0.088 and 0.079, for Brazilian and Uruguayan populations, respectively), and Bayesian cluster analysis inferred two genetic clusters correlated with the geographical origin of isolates. A principal coordinates analysis scatter plot and an unweighted pair group method with arithmetic mean-based dendrogram also grouped Brazilian and Uruguayan isolates into two groups. By pairwise comparison of nitrate-nonutilizing (nit) mutants with a proposed set of testers, all Uruguayan isolates were grouped into a unique vegetative compatibility group (namely VCG 1), while Brazilian isolates were grouped into four VCGs (VCG 1 to 4). Brazilian and Uruguayan populations of C. fructicola were found to be genetically distinct. Our results suggest that isolates of C. fructicola from Brazil capable of causing GLS and ABR arose independently of those from Uruguay. Possible causes leading to the evolutionary differences between populations are discussed.
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Colletotrichum/genética , Malus/microbiologia , Colletotrichum/patogenicidade , Variação Genética , Fenótipo , Phyllachorales , FilogeniaRESUMO
Grapevine fungal trunk diseases (GTDs) have become a serious problem for grapevines worldwide. Nursery vines infected during the propagation process are considered one of the main ways of dissemination of GTD pathogens. In this study, we examined the status of GTDs in grapevine planting material, from rootstocks and scion mother cuttings to grafted rooted vines ready to plant, according to the local nursery propagation process. During 2018-2019, internal symptoms of GTDs were examined in 2400 propagation materials and fungal isolations were carried out from a subsample of 1026 selected materials. Our results revealed that nursery grapevine plants produced in Uruguay have a high incidence of GTDs, regardless of the scion/rootstock combination. Typical brown to black streaks and sectorial wood necrosis were observed in materials on all propagation stages, with a markedly increasing incidence throughout the nursery process, reaching almost 100% in grafted rooted vines ready to plant. Botryosphaeria dieback, Petri disease and black-foot disease were the main GTDs found. The results showed that Botryosphaeria dieback and Petri disease pathogens infect materials from the early stages of the process, with a marked increase towards the end of the plant production process, whereas black-foot disease pathogens were found exclusively in vines ready to plant. Diaporthe dieback pathogens were also detected in materials in all stages but in a low proportion (less than 10% of infected material). Based on single locus analysis, the 180 isolates selected were placed into eight genera and 89% identified within 22 fungal species associated with GTDs, with Phaeoacremonium oleae and Diaporthe terebinthifolii as new records on grapevine worldwide. Our results have concluded that locally produced vines are one of the main ways of dissemination of GTD pathogens and showed that a nursery sanitation programme is required to reduce the incidence of these diseases.
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Plant roots support complex microbial communities that can influence nutrition, plant growth, and health. In grapevine, little is known about the impact of abiotic stresses on the belowground microbiome. In this study, we examined the drought-induced shifts in fungal composition in the root endosphere, the rhizosphere and bulk soil by internal transcribed spacer (ITS) high-throughput amplicon sequencing (HTAS). We imposed three irrigation regimes (100%, 50%, and 25% of the field capacity) to one-year old grapevine rootstock plants cv. SO4 when plants had developed 2-3 roots. Root endosphere, rhizosphere, and bulk soil samples were collected 6- and 12-months post-plantation. Drought significantly modified the overall fungal composition of all three compartments, with the root endosphere compartment showing the greatest divergence from well-watered control (100%). The overall response of the fungal microbiota associated with black-foot disease (Dactylonectria and "Cylindrocarpon" genera) and the potential biocontrol agent Trichoderma to drought stress was consistent across compartments, namely that their relative abundances were significantly higher at 50-100% than at 25% irrigation regime. We identified a significant enrichment in several fungal genera such as the arbuscular mycorrhizal fungus Funneliformis during drought at 25% watering regime within the roots. Our results reveal that drought stress, in addition to its well-characterized effects on plant physiology, also results in the restructuring of grapevine root microbial communities, and suggest the possibility that members of the altered grapevine microbiota might contribute to plant survival under extreme environmental conditions.
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Glomerella leaf spot (GLS) caused by Colletotrichum spp. is a destructive disease of apple restricted to a few regions worldwide. The distribution and evolution of GLS symptoms were observed for two years in Uruguay. The recurrent ascopore production on leaves and the widespread randomized distribution of symptoms throughout trees and orchard, suggest that ascospores play an important role in the disease dispersion. The ability of ascospores to produce typical GLS symptom was demonstrated by artificial inoculation. Colletotrichum strains causing GLS did not result in rot development, despite remaining alive in fruit lesions. Based on phylogenetic analysis of actin, ß-tubulin and glyceraldehyde-3-phosphate dehydrogenase gene regions of 46 isolates, 25 from fruits and 21 from leaves, C. karstii was identified for the first time causing GLS in Uruguay and C. fructicola was found to be the most frequent (89%) and aggressive species. The higher aggressiveness of C. fructicola and its ability on to produce abundant fertile perithecia could help to explain the predominance of this species in the field.
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INTRODUCTION: In recent decades, the prevalence of carbapenem-resistant Acinetobacter isolates has increased, and the production of oxacillinase (OXA)-type carbapenemases is the main mechanism underlying resistance. We evaluated OXA production from 114 Acinetobacter isolates collected between March and December 2013 from different clinical specimens of patients in two hospitals (Hospital 1 [n = 61] and Hospital 2 [n = 53]) located in Niterói, Rio de Janeiro, Brazil. We also evaluated the genetic diversity of OXA-producing isolates. METHODS: All the isolates were identified through the automated system Vitek II and matrix-assisted laser desorption ionization-time of flight mass spectrometry MALDI-TOF MS as belonging to the A. baumannii-A. calcoaceticuscomplex. Antimicrobial susceptibility profiles were verified through agar diffusion tests. The presence of OXA-encoding genes was confirmed by PCR. The genetic diversity of isolates positive for carbapenemase production was analyzed through pulsed-field gel electrophoresis. RESULTS: There was a high rate of resistance to carbapenems in the isolates (imipenem: 96%; meropenem: 92%) from both hospitals. Moreover, a high percentage (95.6%) of OXA-23-positive isolates was observed for both hospitals, indicating that this was the main mechanism of carbapenem-resistance among the studied population. In addition, most isolates (96.5%) were positive for bla OXA-51. A high genetic diversity and a few major genotypes were found among the OXA-23-positive isolates analyzed. Only intra-hospital dissemination was observed. CONCLUSIONS: The elevated dissemination of bla OXA-23-like observed among Acinetobacter isolates from both the studied hospitals highlights the need for continuous epidemiological surveillance in these institutions.
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Infecções por Acinetobacter/microbiologia , Acinetobacter/enzimologia , beta-Lactamases/biossíntese , Acinetobacter/efeitos dos fármacos , Acinetobacter/genética , Antibacterianos/farmacologia , Técnicas de Tipagem Bacteriana , Brasil , DNA Bacteriano/genética , Eletroforese em Gel de Campo Pulsado , Hospitais Gerais , Humanos , Testes de Sensibilidade Microbiana , Reação em Cadeia da Polimerase , beta-Lactamases/efeitos dos fármacosRESUMO
Colletotrichum fructicola causes two important diseases on apple in Southern Brazil, bitter rot (ABR) and Glomerella leaf spot (GLS). In this pathosystem, the Colletotrichum ability to cause different symptoms could be related to differences of extracellular enzymes produced by the fungi. Thus, the objectives of this study were to compare the production of these enzymes between ABR- and GLS-isolate in vitro and to evaluate their involvement on infected apple leaves with C. fructicola. In agar plate enzymatic assay, ABR- showed significantly higher amylolytic and pectolytic activity than GLS-isolate. In contrast, for lipolytic and proteolytic no significant differences were observed between isolates. In culture broth, ABR-isolate also had higher activity of pectin lyase (PNL), polygalacturonase (PG) and laccase (LAC). Notably, LAC was significantly five-fold higher in ABR- than GLS-isolate. On the other hand, in infected apple leaves no significant difference was observed between isolates for PNL, PG and LAC. Although differences in extracellular enzymes of ABR- and GLS-isolate have not been observed in vivo, these results contributed to highlight the importance to investigate such enzymes in depth.
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Apple bitter rot (ABR) and Glomerella leaf spot (GLS) can be caused by Colletotrichum fructicola. Although both diseases can occur simultaneously in orchards, some isolates show clear organ specialization. Thus, this work was aimed to compare microscopically the development of preinfective structures of ABR- and GLS isolates and their impact on the enzymatic oxidant defense system during the leaf infection process. On leaves, conidial germlings of GLS-isolate formed appressoria mostly sessile. In contrast, those of ABR-isolate were pedicellate and formed multiple melanized appressoria probably as a sign of unsuccessful infection attempts. Neither ABR- nor GLS isolate triggered hypersensitive response in apple leaves. In overall, the activity of scavenging enzymes was higher and long-lasting in leaves inoculated by GLS- than by ABR isolate and control. Guaiacol peroxidase, catalase, and glutathione reductase had activity peaks within 24 h after inoculation (HAI). Ascorbate peroxidase activity was higher only in GLS-infected leaves at 6 HAI, while superoxide dismutase remained unaltered. A lower level of hydrogen peroxide (H2O2) was determined in GLS-infected plants at 48 HAI, but the electrolyte leakage markedly increased. Disease symptoms in leaves were only caused by GLS-isolate. Results suggest that the virulent isolate coordinately downregulates the oxidative plant defense responses enabling its successful establishment in apple leaves.
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Colletotrichum/isolamento & purificação , Malus/microbiologia , Doenças das Plantas/microbiologia , Colletotrichum/genética , Colletotrichum/crescimento & desenvolvimento , Colletotrichum/metabolismo , Peróxido de Hidrogênio , Malus/enzimologia , Malus/genética , Malus/metabolismo , Estresse Oxidativo , Peroxidase/genética , Peroxidase/metabolismo , Doenças das Plantas/genética , Folhas de Planta/enzimologia , Folhas de Planta/genética , Folhas de Planta/metabolismo , Folhas de Planta/microbiologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Esporos Fúngicos/classificação , Esporos Fúngicos/genética , Esporos Fúngicos/crescimento & desenvolvimento , Esporos Fúngicos/isolamento & purificação , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismoRESUMO
Colletotrichum species are associated with Apple bitter rot (ABR) and Glomerella leaf spot (GLS). Whereas both apple diseases occur frequently in Brazil, only the former has been reported in Uruguay. This work was aimed at identifying and comparing morpho-cultural characteristics and pathogenic variability of thirty-nine Colletotrichum isolates from both countries. Sequencing of the internal transcribed spacer (ITS) rDNA, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and ß-tubulin (TUB2) allowed the identification of three species causing ABR and GLS in Brazil, i.e., Colletotrichum fructicola, Colletotrichum karstii, and Colletotrichum nymphaeae; and three species causing ABR in Uruguay, i.e., C. fructicola, Colletotrichum theobromicola, and Colletotrichum melonis. Six groups of colony colours were recorded with group 1 (mycelium white to pink and in reverse pinkish) and group 2 (mycelium white to grey and in reverse pinkish) the most frequent. Isolates of C. fructicola and C. theobromicola were sensitive to benomyl, while C. karstii, C. nymphaeae, and C. melonis were resistant. Conidia were predominantly cylindrical for C. fructicola and C. karstii, fusiform for C. nymphaeae and C. melonis, and obclavate for C. theobromicola. Brazilian isolates caused ABR in wounded fruits, but only five in non-wounded ones. Uruguayan isolates produced symptoms in fruits with or without previous wounding. All Brazilian isolates from GLS and twelve from ABR were able to cause GLS symptoms, while a sole Uruguayan ABR-isolate caused leaf spot symptoms. This study gives a better insight on the new species causing apple disease in both countries and discusses their pathogenic potential.
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Colletotrichum/classificação , Colletotrichum/isolamento & purificação , Malus/microbiologia , Doenças das Plantas/microbiologia , Brasil , Análise por Conglomerados , Colletotrichum/citologia , Colletotrichum/genética , DNA Espaçador Ribossômico/química , DNA Espaçador Ribossômico/genética , Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora)/genética , Dados de Sequência Molecular , Micélio/citologia , Filogenia , Análise de Sequência de DNA , Esporos Fúngicos/citologia , Tubulina (Proteína)/genética , UruguaiRESUMO
INTRODUCTION: Acquired production of metallo-ß-lactamases is an important mechanism of resistance in Pseudomonas aeruginosa. The objective of this study was to investigate the production of metallo-ß-lactamase and the genetic diversity among ceftazidime-resistant P. aeruginosa isolates from State of Sergipe, Brazil. METHODS: Metallo-ß-lactamase was investigated using the disk approximation test and polymerase chain reaction (PCR). Genetic diversity was evaluated by pulsed-field gel electrophoresis (PFGE). RESULTS: A total of 48 (51.6%) isolates were resistant to ceftazidime. Six (12.2%) of these were positive for metallo-ß-lactamase production. Only two (4.1%) of the ceftazidime-resistant isolates carried the bla SPM-1 gene. CONCLUSIONS: Production of metallo-ß-lactamases was not the main mechanism of resistance to ceftazidime and carbapenems among P. aeruginosa strains in Sergipe, Brazil.
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Pseudomonas aeruginosa/enzimologia , beta-Lactamases/biossíntese , Antibacterianos/farmacologia , Brasil , Testes de Sensibilidade a Antimicrobianos por Disco-Difusão , Farmacorresistência Bacteriana , Eletroforese em Gel de Campo Pulsado , Variação Genética , Humanos , Reação em Cadeia da Polimerase , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/genéticaRESUMO
CONTEXT AND OBJECTIVES: Urinary tract infections are the most common cause of hospital-acquired infections, and the use of indwelling urinary catheters is a predisposing factor for their development. The aims of this study were to estimate the frequency of pre and postoperative bacteriuria, identify the microorganisms involved, count the colony-forming units, determine the antibiotic sensitivity profile and compare the results from pre and postoperative urinalyses among women undergoing gynecological surgery with implantation of a urinary catheter. DESIGN AND SETTING: Non-controlled prospective observational single-cohort epidemiological study carried out at a university hospital. METHODS: Urine samples were collected before and 24 hours after catheterization for urinalysis, culturing and antibiotic sensitivity testing. Pre and postoperative urinalyses were compared using Wilcoxon and McNemar non-parametric tests. RESULTS: Fifty-one women participated in the study. Escherichia coligrew in six preoperative samples (11.8%) and Klebsiella pneumoniae in one (1.9%), but bacterial growth did not occur in any postoperative sample. Urinalysis showed lower number of pus cells in the postoperative urine samples (P < 0.05). There were no differences in red blood cell counts or in the nitrite and leukocyte esterase tests, between the samples. CONCLUSION: Bacteriuria was found in 13.7% of the preoperative samples. Gram-negative bacteria sensitive to most antibiotics were identified. In the postoperative samples, no bacterial growth was observed. Urinalysis only showed significant reduction of leukocyturia in the postoperative period.
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Antibacterianos/uso terapêutico , Antibioticoprofilaxia/métodos , Bacteriúria/microbiologia , Bacteriúria/prevenção & controle , Procedimentos Cirúrgicos em Ginecologia/efeitos adversos , Cateterismo Urinário/efeitos adversos , Cateteres Urinários/efeitos adversos , Adolescente , Adulto , Idoso , Contagem de Colônia Microbiana , Infecção Hospitalar/microbiologia , Escherichia coli/isolamento & purificação , Feminino , Humanos , Klebsiella/isolamento & purificação , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Período Pós-Operatório , Período Pré-Operatório , Estudos Prospectivos , Estatísticas não Paramétricas , Fatores de Tempo , Urinálise , Adulto JovemRESUMO
The phenotypic and genotypic characteristics of clinical and intestinal enterococcal isolates recovered from inpatients and outpatients of two Brazilian hospitals, located in Niterói city, Rio de Janeiro, Brazil, were compared. A total of 601 strains were studied, including 253 isolated from different clinical sources and 348 intestinal strains (205 isolated from inpatients and 143 from outpatients) recovered from fecal specimens. Isolates were identified by using conventional physiological tests and evaluated for high-level resistance to aminoglycosides (HLR-A) and resistance to vancomycin and ampicillin by the agar screening technique. Susceptibility to several antimicrobial agents was evaluated by the disk diffusion method. The genetic diversity of Enterococcus faecalis strains presenting HLR-A was assessed by pulsed-field gel electrophoresis of chromosomal DNA after SmaI digestion. E. faecalis was the most frequent species among clinical isolates (90.1%) and intestinal strains from inpatients (53.6%). E. casseliflavus was the prevalent species among intestinal isolates from outpatients (35.0%). Clinical isolates were shown to be resistant to erythromycin (53.0%), tetracycline (52.2%), ciprofloxacin (36.4%), gentamicin (36.4%), streptomycin (30.4%), chloramphenicol (34.4%), norfloxacin (32.0%), imipenem (3.2%), and ampicillin (2.8%). Vancomycin resistance was only detected in intrinsic vancomycin-resistant enterococcal species. The overall prevalence of HLR-A was 52.2% among clinical isolates and 40.5% among intestinal strains. However, HLR-A was significantly more frequent among intestinal strains obtained from inpatients (56.6%) than among strains from outpatients (17.5%). Three major clonal groups were found among E. faecalis strains exhibiting HLR-GE or HLR-GE/ST (clonal groups GE-A and GE-B), and strains exhibiting HLR-ST (clonal group ST-A). HLR-A, particularly HLR-GE, was most frequently associated with enterococcal strains of nosocomial origin. Isolates included in the major clonal groups were recovered from clinical and intestinal sources from patients in both hospitals, indicating both intrahospital and interhospital spread of strains.
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Infecção Hospitalar/microbiologia , Enterococcus/efeitos dos fármacos , Enterococcus/genética , Infecções por Bactérias Gram-Negativas/microbiologia , Intestinos/microbiologia , Brasil/epidemiologia , Cromossomos Bacterianos/genética , Farmacorresistência Bacteriana , Eletroforese em Gel de Campo Pulsado , Fezes/microbiologia , Genótipo , Infecções por Bactérias Gram-Negativas/epidemiologia , Humanos , Pacientes Internados , Testes de Sensibilidade Microbiana , Pacientes Ambulatoriais , Fenótipo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Interpretation of the mupirocin E-test for low-level mupirocin-resistant Staphylococcus aureus strains has been improved by adding the indicator dye tetrazolium. E-tests were compared with agar dilution methods for assessing mupirocin susceptibility. MICs obtained by the agar dilution method and E-tests showed 89.3% agreement within 2 log(2) dilution criteria. The agreement between MICs increased to 100% in the 1 log(2) dilution definition when the indicator dye tetrazolium was added to the E-test. The use of the E-test with tetrazolium reduction is more accurate for determining mupirocin MICs for S. aureus strains.
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Antibacterianos/farmacologia , Testes de Sensibilidade Microbiana/normas , Mupirocina/farmacologia , Staphylococcus aureus/efeitos dos fármacos , Infecção Hospitalar/microbiologia , Farmacorresistência Bacteriana , Humanos , Indicadores e Reagentes/química , Testes de Sensibilidade Microbiana/métodos , Nitroazul de Tetrazólio/química , Variações Dependentes do Observador , Reprodutibilidade dos Testes , Infecções Estafilocócicas/microbiologiaRESUMO
Abstract INTRODUCTION In recent decades, the prevalence of carbapenem-resistant Acinetobacter isolates has increased, and the production of oxacillinase (OXA)-type carbapenemases is the main mechanism underlying resistance. We evaluated OXA production from 114 Acinetobacter isolates collected between March and December 2013 from different clinical specimens of patients in two hospitals (Hospital 1 [n = 61] and Hospital 2 [n = 53]) located in Niterói, Rio de Janeiro, Brazil. We also evaluated the genetic diversity of OXA-producing isolates. METHODS All the isolates were identified through the automated system Vitek II and matrix-assisted laser desorption ionization-time of flight mass spectrometry MALDI-TOF MS as belonging to the A. baumannii-A. calcoaceticuscomplex. Antimicrobial susceptibility profiles were verified through agar diffusion tests. The presence of OXA-encoding genes was confirmed by PCR. The genetic diversity of isolates positive for carbapenemase production was analyzed through pulsed-field gel electrophoresis. RESULTS There was a high rate of resistance to carbapenems in the isolates (imipenem: 96%; meropenem: 92%) from both hospitals. Moreover, a high percentage (95.6%) of OXA-23-positive isolates was observed for both hospitals, indicating that this was the main mechanism of carbapenem-resistance among the studied population. In addition, most isolates (96.5%) were positive for bla OXA-51. A high genetic diversity and a few major genotypes were found among the OXA-23-positive isolates analyzed. Only intra-hospital dissemination was observed. CONCLUSIONS The elevated dissemination of bla OXA-23-like observed among Acinetobacter isolates from both the studied hospitals highlights the need for continuous epidemiological surveillance in these institutions.
Assuntos
Humanos , Acinetobacter/enzimologia , beta-Lactamases/efeitos dos fármacos , Infecções por Acinetobacter/microbiologia , Acinetobacter/efeitos dos fármacos , Acinetobacter/genética , beta-Lactamases/biossíntese , Brasil , DNA Bacteriano/genética , Testes de Sensibilidade Microbiana , Reação em Cadeia da Polimerase , Técnicas de Tipagem Bacteriana , Eletroforese em Gel de Campo Pulsado , Hospitais Gerais , Antibacterianos/farmacologiaRESUMO
INTRODUCTION: Acinetobacter spp. have emerged as notorious pathogens involved in healthcare-associated infections. Carbapenems are important antimicrobial agents for treating infections due to multidrug resistant Acinetobacter spp. Different mechanisms may confer resistance to these drugs in the genus, particularly production of class D carbapenemases. OXA-23-like family has been pointed out as one of the predominant carbapenamases among Acinetobacter. The present work aimed to investigate the occurrence of OXA-23-like carbapenemases among Acinetobacter isolates recovered from patients of a university hospital in Niterói, RJ, Brazil. METHODS: Antimicrobial susceptibility profiles were determined by disk-diffusion. Imipenem resistant isolates were submitted to Modified Hodge Test in order to screen for carbapenemase production, and later to polymerase chain reaction (PCR) to investigate the presence of bla(OXA-23). RESULTS: Imipenem and meropenem resistance rates were 71.4% and 69.7%, respectively. The Modified Hodge Test revealed carbapenemase production among 76 (89.4%) of the 85 imipenem resistant isolates analyzed; according to PCR results, 81 isolates (95.4%) carried the bla(OXA-23) gene. CONCLUSIONS: OXA-23-like enzymes may be an important mechanism of carbapenem resistance among isolates present in the hospital studied.
Assuntos
Acinetobacter/enzimologia , beta-Lactamases/análise , Acinetobacter/efeitos dos fármacos , Acinetobacter/isolamento & purificação , Antibacterianos/farmacologia , Brasil , Testes de Sensibilidade a Antimicrobianos por Disco-Difusão , Farmacorresistência Bacteriana , Hospitais Universitários , Humanos , Testes de Sensibilidade Microbiana , Reação em Cadeia da Polimerase , beta-Lactamases/genéticaRESUMO
ABSTRACT CONTEXT AND OBJECTIVES: Urinary tract infections are the most common cause of hospital-acquired infections, and the use of indwelling urinary catheters is a predisposing factor for their development. The aims of this study were to estimate the frequency of pre and postoperative bacteriuria, identify the microorganisms involved, count the colony-forming units, determine the antibiotic sensitivity profile and compare the results from pre and postoperative urinalyses among women undergoing gynecological surgery with implantation of a urinary catheter. DESIGN AND SETTING: Non-controlled prospective observational single-cohort epidemiological study carried out at a university hospital. METHODS: Urine samples were collected before and 24 hours after catheterization for urinalysis, culturing and antibiotic sensitivity testing. Pre and postoperative urinalyses were compared using Wilcoxon and McNemar non-parametric tests. RESULTS: Fifty-one women participated in the study. Escherichia coligrew in six preoperative samples (11.8%) and Klebsiella pneumoniae in one (1.9%), but bacterial growth did not occur in any postoperative sample. Urinalysis showed lower number of pus cells in the postoperative urine samples (P < 0.05). There were no differences in red blood cell counts or in the nitrite and leukocyte esterase tests, between the samples. CONCLUSION: Bacteriuria was found in 13.7% of the preoperative samples. Gram-negative bacteria sensitive to most antibiotics were identified. In the postoperative samples, no bacterial growth was observed. Urinalysis only showed significant reduction of leukocyturia in the postoperative period.
RESUMO CONTEXTO E OBJETIVOS: As infecções urinárias são a causa mais comum de infecções hospitalares, e o uso de cateteres de demora é fator predisponente para o seu desenvolvimento. Os objetivos deste estudo foram estimar a frequência de bacteriúria pré e pós-operatória, identificar os germes encontrados, a contagem de unidades formadoras de colônias e o perfil de sensibilidade aos antibióticos, além de comparar os resultados dos exames de urina pré- e pós-operatórios em mulheres submetidas a cirurgias ginecológicas com cateterismo vesical. TIPO DE ESTUDO E LOCAL: Estudo epidemiológico, observacional, de coorte única, prospectivo, não controlado, realizado em hospital universitário. MÉTODOS: Amostras de urina foram colhidas antes da cateterização e após 24 horas para urinálise, cultura e antibiograma. Os resultados da urinálise no pré- e pós-operatório foram comparados utilizando-se os testes não paramétricos de Wilcoxon e McNemar. RESULTADOS: Participaram do estudo 51 mulheres. Houve crescimento de Escherichia coli em seis amostras pré-operatórias (11,8%) e deKlebsiella pneumoniae em uma (1,9%), mas não houve crescimento bacteriano em nenhuma amostra pós-operatória. A urinálise mostrou menor quantidade de piócitos na amostra de urina pós-cirúrgica (P < 0,05). Não houve diferença quanto ao número de hemácias e às reações para nitrito e leucocitesterase, entre as amostras. CONCLUSÃO: Houve bacteriúria em 13,7% das amostras pré-operatórias, sendo identificadas bactérias Gram-negativas sensíveis à maioria dos antibióticos. Não foi observado crescimento bacteriano nas amostras pós-operatórias. A urinálise mostrou somente redução significativa da leucocitúria no pós-operatório.
Assuntos
Adolescente , Adulto , Idoso , Feminino , Humanos , Pessoa de Meia-Idade , Adulto Jovem , Antibacterianos/uso terapêutico , Antibioticoprofilaxia/métodos , Bacteriúria/microbiologia , Bacteriúria/prevenção & controle , Procedimentos Cirúrgicos em Ginecologia/efeitos adversos , Cateterismo Urinário/efeitos adversos , Cateteres Urinários/efeitos adversos , Contagem de Colônia Microbiana , Infecção Hospitalar/microbiologia , Escherichia coli/isolamento & purificação , Klebsiella/isolamento & purificação , Testes de Sensibilidade Microbiana , Período Pós-Operatório , Período Pré-Operatório , Estudos Prospectivos , Estatísticas não Paramétricas , Fatores de Tempo , UrináliseRESUMO
We report the case of a patient with chronic obstructive pulmonary disease and bronchiectasis, chronically using corticosteroids, who acquired pulmonary nocardiosis, which presented as multiple cavitated nodules. The principal symptoms were fever, dyspnea and productive cough with purulent sputum. Chest X-ray and computed tomography of the chest revealed nodules, some of which were cavitated, in both lungs. Sputum smear microscopy and culture revealed the presence of Nocardia spp. The patient was treated with imipenem and cilastatin, which produced an excellent clinical response.