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Emergence of novel mutations of caspofungin resistance in candida glabrata with prolonged therapy
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PATHOLOGY UPDATE 2010 ABSTRACTS
Discussion: A correlation between WT1 expression levels and known risk factors for early relapse is demonstrated. In the minimal residual disease setting, detectable WT1 levels in peripheral blood post-induction and post-consolidation predicted very poor leukaemia free survival.
CARCINOGENIC EFFECT OF THE NSAID SULINDAC IN THE MOUSE PROXIMAL COLON Ruta Gupta1, Dessislava Mladenova2, Jane E. Dahlstrom1, Elaine Bean1, Maija Kohonen-Corish2 1 ACT Pathology, The Canberra Hospital and Australian National University Medical School, ACT and 2Cancer Research Program, Garvan Institute of Medical Research, Sydney, NSW, Australia Background: Procarcinogenic effects of the non-steroidal anti-inflammatory drug (NSAID) sulindac have been described in the mouse proximal colon. The possible role of hypoxia inducible factors (HIF) in this process is poorly understood. Aim: To study histological effects of sulindac in HIF1a knockout mice. Methods: Mice were bred with a specific homozygous HIF1a deletion in the colonic epithelium (n ¼ 16) with their genotype controls that express HIF1a in the colon (n ¼ 19). They were given a diet containing 320 ppm sulindac for 20 weeks and the dietary controls (n ¼ 17) received mouse chow without sulindac. Biopsies from proximal to distal mouse colons were examined by light microscopy. The average number of biopsies analysed per mouse was 7.9 for the test and 8.5 for the control genotype. SPSS (Version 11) and StatXact 8 software were used for statistical analysis. Results: Sulindac exposure was associated with acute and chronic inflammation. Neovascularisation, apoptosis, necrosis and fibrosis were not present. Neoplastic transformation was only seen in the mice receiving the sulindac diet (p 5 0.0001). There was less colon inflammation in the HIF1a knockout mice compared with the genotype controls (p ¼ 0.004). Conclusion: HIF1a may have a pro-inflammatory role in sulindac-induced carcinogenesis in the mouse proximal colon.
EMERGENCE OF NOVEL MUTATIONS OF CASPOFUNGIN RESISTANCE IN CANDIDA GLABRATA WITH PROLONGED THERAPY Marjoree M. Sehu, Hanna Sidjabat, David Ellis, Joe McCormack, E. Geoffrey Playford, David L. Paterson Pathology Queensland, Princess Alexandra Hospital, Woolloongabba, Queensland, Australia Background: Resistance to echinocandin is rare but resistance emerging during therapy has been reported in the literature. In Candida glabrata, mutations in the FKS1 and FKS2 regions of the glucan synthase complex has been reported.
Pathology (2010), 42(S1)
Two clinical isolates of Candida glabrata from Queensland were noted to be caspofungin resistant after prolonged caspofungin therapy. Aim: The aim of this study is to look at the resistance mutation in the Candida glabrata isolates. Method: The caspofungin minimum inhibitory concentration (MIC) for all isolates was confirmed using CLSI M27-A2 methodology. Isogenicity of the isolates was verified by multilocus sequence typing (MLST). DNA was extracted from the isolates and the regions of interest of the glucan synthase enzyme complex were then sequenced against wild type sequences. Results: The raised MIC to caspofungin of the clinical cases was confirmed. Isogenicity was verified with the presence of a persistent unique strain. Two novel mutations in the FKS2 glucan synthase enxyme complex were identified on sequencing, which has not been previously reported. Discussion: Candidaemia by non-Candida albicans is increasing. Resistance to caspofungin may lead to treatment failure. It also leaves few alternatives for treatment of lifethreatening candidiasis. Emerging resistance to caspofungin in C. glabrata, especially in cases of prolonged caspofungin exposure, creates the need for increased surveillance and heightened suspicion in the microbiology laboratory as well as in the clinical setting.
VALIDATION OF CALCULATED IONISED CALCIUM FOR CRITICALLY ILL PATIENTS S. D. C. Thomas, B. D. Rumbelow, G. H. White Chemical Pathology, SA Pathology, Flinders Medical Centre, Adelaide, South Australia Aim: A previously published formula calculated plasma ionised calcium (iCa) from several plasma variables and was validated for healthy subjects. This study assessed the validity of the formula in critically ill patients. Methods: Plasma iCa calculated using plasma albumin, total protein (for calculation of globulins), bicarbonate and total calcium concentrations measured on an automated analyser in routine samples of 365 critically ill patients, was compared with iCa measured with an ion selective electrode. Results: The calculated and measured iCa ranges were 0.86–1.77 mmol/L and 0.87–1.72 mmol/L, respectively (reference interval 1.17–1.31 mmol/L). The bias between the methods was 0.004 mmol/L (Bland Altman). Linear regression between the values showed a high correlation [r2 ¼ 0.90; calculated iCa ¼ (0.902.measured iCa) þ 0.1299]. For patients with albumin 5 20 g/L the bias was 0.005 mmol/L [r2 ¼ 0.95; calculated iCa ¼ (1.0792.measured iCa) þ 0.091]. Critical differences at medical decision points were acceptable. Discussion: Although calculated iCa has been used in many clinical situations, agreement between the methods has not been shown in critically ill patients. Calculated ionised calcium can be used to assess calcium status in critically ill patients, including those with low albumin.
PATHOLOGY UPDATE 2010 ABSTRACTS
Discussion: A correlation between WT1 expression levels and known risk factors for early relapse is demonstrated. In the minimal residual disease setting, detectable WT1 levels in peripheral blood post-induction and post-consolidation predicted very poor leukaemia free survival.
CARCINOGENIC EFFECT OF THE NSAID SULINDAC IN THE MOUSE PROXIMAL COLON Ruta Gupta1, Dessislava Mladenova2, Jane E. Dahlstrom1, Elaine Bean1, Maija Kohonen-Corish2 1 ACT Pathology, The Canberra Hospital and Australian National University Medical School, ACT and 2Cancer Research Program, Garvan Institute of Medical Research, Sydney, NSW, Australia Background: Procarcinogenic effects of the non-steroidal anti-inflammatory drug (NSAID) sulindac have been described in the mouse proximal colon. The possible role of hypoxia inducible factors (HIF) in this process is poorly understood. Aim: To study histological effects of sulindac in HIF1a knockout mice. Methods: Mice were bred with a specific homozygous HIF1a deletion in the colonic epithelium (n ¼ 16) with their genotype controls that express HIF1a in the colon (n ¼ 19). They were given a diet containing 320 ppm sulindac for 20 weeks and the dietary controls (n ¼ 17) received mouse chow without sulindac. Biopsies from proximal to distal mouse colons were examined by light microscopy. The average number of biopsies analysed per mouse was 7.9 for the test and 8.5 for the control genotype. SPSS (Version 11) and StatXact 8 software were used for statistical analysis. Results: Sulindac exposure was associated with acute and chronic inflammation. Neovascularisation, apoptosis, necrosis and fibrosis were not present. Neoplastic transformation was only seen in the mice receiving the sulindac diet (p 5 0.0001). There was less colon inflammation in the HIF1a knockout mice compared with the genotype controls (p ¼ 0.004). Conclusion: HIF1a may have a pro-inflammatory role in sulindac-induced carcinogenesis in the mouse proximal colon.
EMERGENCE OF NOVEL MUTATIONS OF CASPOFUNGIN RESISTANCE IN CANDIDA GLABRATA WITH PROLONGED THERAPY Marjoree M. Sehu, Hanna Sidjabat, David Ellis, Joe McCormack, E. Geoffrey Playford, David L. Paterson Pathology Queensland, Princess Alexandra Hospital, Woolloongabba, Queensland, Australia Background: Resistance to echinocandin is rare but resistance emerging during therapy has been reported in the literature. In Candida glabrata, mutations in the FKS1 and FKS2 regions of the glucan synthase complex has been reported.
Pathology (2010), 42(S1)
Two clinical isolates of Candida glabrata from Queensland were noted to be caspofungin resistant after prolonged caspofungin therapy. Aim: The aim of this study is to look at the resistance mutation in the Candida glabrata isolates. Method: The caspofungin minimum inhibitory concentration (MIC) for all isolates was confirmed using CLSI M27-A2 methodology. Isogenicity of the isolates was verified by multilocus sequence typing (MLST). DNA was extracted from the isolates and the regions of interest of the glucan synthase enzyme complex were then sequenced against wild type sequences. Results: The raised MIC to caspofungin of the clinical cases was confirmed. Isogenicity was verified with the presence of a persistent unique strain. Two novel mutations in the FKS2 glucan synthase enxyme complex were identified on sequencing, which has not been previously reported. Discussion: Candidaemia by non-Candida albicans is increasing. Resistance to caspofungin may lead to treatment failure. It also leaves few alternatives for treatment of lifethreatening candidiasis. Emerging resistance to caspofungin in C. glabrata, especially in cases of prolonged caspofungin exposure, creates the need for increased surveillance and heightened suspicion in the microbiology laboratory as well as in the clinical setting.
VALIDATION OF CALCULATED IONISED CALCIUM FOR CRITICALLY ILL PATIENTS S. D. C. Thomas, B. D. Rumbelow, G. H. White Chemical Pathology, SA Pathology, Flinders Medical Centre, Adelaide, South Australia Aim: A previously published formula calculated plasma ionised calcium (iCa) from several plasma variables and was validated for healthy subjects. This study assessed the validity of the formula in critically ill patients. Methods: Plasma iCa calculated using plasma albumin, total protein (for calculation of globulins), bicarbonate and total calcium concentrations measured on an automated analyser in routine samples of 365 critically ill patients, was compared with iCa measured with an ion selective electrode. Results: The calculated and measured iCa ranges were 0.86–1.77 mmol/L and 0.87–1.72 mmol/L, respectively (reference interval 1.17–1.31 mmol/L). The bias between the methods was 0.004 mmol/L (Bland Altman). Linear regression between the values showed a high correlation [r2 ¼ 0.90; calculated iCa ¼ (0.902.measured iCa) þ 0.1299]. For patients with albumin 5 20 g/L the bias was 0.005 mmol/L [r2 ¼ 0.95; calculated iCa ¼ (1.0792.measured iCa) þ 0.091]. Critical differences at medical decision points were acceptable. Discussion: Although calculated iCa has been used in many clinical situations, agreement between the methods has not been shown in critically ill patients. Calculated ionised calcium can be used to assess calcium status in critically ill patients, including those with low albumin.