VIM Genes Research Articles

Aphthous ulcer drug inhibits prostate tumor metastasis by targeting IKKɛ/TBK1/NF-κB signaling.

Tumor metastasis is the major cause of death for prostate cancer (PCa) patients. However, the treatment options for metastatic PCa are very limited. Epithelial-mesenchymal transition (EMT) has been reported to be an indispensable step for tumor metastasis and is suggested to associate with acquisition of cancer stem cell (CSC) attributes. We propose that small-molecule compounds that can reverse EMT or induce mesenchymal-epithelial transition (MET) of PCa cells may serve as drug candidates for anti-metastasis therapy.Methods: The promoters of CDH1 and VIM genes were sub-cloned to drive the expression of firefly and renilla luciferase reporter in a lentiviral vector. Mesenchymal-like PCa cells were infected with the luciferase reporter lentivirus and subjected to drug screening from a 1274 approved small-molecule drug library for the identification of agents to reverse EMT. The dosage-dependent effect of candidate compounds was confirmed by luciferase reporter assay and immunoblotting. Wound-healing assay, sphere formation, transwell migration assay, and in vivo intracardiac and orthotopic tumor xenograft experiments were used to evaluate the mobility, metastasis and tumor initiating capacity of PCa cells upon treatment. Possible downstream signaling pathways affected by the candidate compound treatment were analyzed by RNA sequencing and immunoblotting.Results: Drug screening identified Amlexanox, a drug used for recurrent aphthous ulcers, as a strong agent to reverse EMT. Amlexanox induced significant suppression of cell mobility, invasion, serial sphere formation and in vivo metastasis and tumor initiating capacity of PCa cells. Amlexanox treatment led to downregulation of the IKK-ɛ/ TBK1/ NF-κB signaling pathway. The effect of Amlexanox on EMT reversion and cell mobility inhibition can be mimicked by other IKK-ɛ/TBK1 inhibitors and rescued by reconstitution of dominant active NF-κB.Conclusions: Amlexanox can sufficiently suppress PCa metastasis by reversing EMT through downregulating the IKK-ɛ/TBK1/NF-κB signaling axis.

Detection of Antibiotic Resistance Determinants and Their Transmissibility among Clinically Isolated Carbapenem-Resistant Escherichia coli from South India

Objectives: The aim of this study was to analyze the prevalence of the CTX-M, TEM, SHV, VIM, NDM, and OXA genes in carbapenemase-producing Escherichia coli and their transmissibility at a tertiary care hospital in south India. Materials and Methods: Twenty-one carbapenem-resistant E. coli (carbapenem-resistant Enterobacteriaceae; CRE) were collected from the Sri Sathya Sai Institute of Higher Medical Sciences (Puttaparthi India). Resistance to antibiotics was analyzed by Vitek-2, and the identity of the isolates was confirmed by 16S rDNA sequencing. RAPD and enterobacterial repetitive intergenic consensus (ERIC)-PCR were performed for molecular typing. Metallo-β-lactamase production was confirmed by a double disc synergy test. The presence of the extended-spectrum β-lactamases CTX-M, TEM, and SHV and of the carbapenemases NDM, VIM, and OXA was determined by PCR. Carbapenemase variants were further confirmed by sequencing. The transmissibility of the genes was tested by conjugation. Results: Twelve of the 21 (57%) carbapenem-resistant E. coli isolates were community acquired, indicating the spread of CRE in environmental samples. TEM and NDM-5 were found to be the major β-lactamases produced by the pathogens. OXA-181 was found in 5 of the isolates. All 21 isolates were found to harbor more than one of the tested β-lactamases, and all of the isolates were found to have the capacity to participate in conjugation; 15 of the transconjugants were found to have acquired the tested β-lactamases, substantiating their ability to be transferred to other strains of bacteria. Conclusion: Monitoring of community-acquired carbapenem-resistant bacteria is very important as the association of resistance determinants with mobile genetic elements would present a serious clinical challenge.

Molecular Characterization of Carbapenemase Producing Klebsiella Pneumoniae Dominance of OXA-48, KPC, VIM and NDM Producers in Khartoum, Sudan

Background and Objectives: Little is known about carbapenemase producing Klebsiella pneumoniae (CPK) in Sudan. This study aimed to determine the prevalence of CPK in a major hospital in Khartoum, Sudan between may 2015 - January 2017 and to characterize the isolates and detect the types of carbapenemase (s) they produced. Materials and Methods: The study was done in the Department of Molecular Microbiology, Faculty of Medical Laboratories Science, Al-Neleen University. All the isolates were obtained from clinical samples of patients treated inside the hospitals. Strains of K. pneumoniae resistant to at least one carbapenem (imipenem or meropenem) by using disc diffusion technique according to the CLSI guidelines were included in this study. Molecular detection of carbapenemase genes was achieved using Real-Time PCR (Sacace Biotechnologie, Italie). Results: A total 96 strains of K. pneumoniae of different non duplicated isolates were obtained from different hospitals. Seventy-two percent (70/96) isolates were positive for carbapenemase genes; 59.4% (57/96) were positive for blaKPC genes, 57.3% (55/96) were positive for blaNDM genes, 37.5% (36/96) were positive for blaVIM genes and 35.4% (34/96) were positive for blaOXA-48 genes. Nineteen isolates possessed four genes (blaKPC, blaNDM, blaVIM and blaOXA-48), fourteen isolates possessed three genes{( blaNDM, blaVIM and blaOXA-48=6), (blaKPC, blaNDM, and blaOXA-48=3), (blaKPC, blaNDM and blaVIM =3), (blaKPC, blaVIM and blaOXA-48=2)}, 27 isolates possessed two genes{( blaKPC and blaNDM =21), (blaKPC, blaOXA-48=2), (blaNDM and blaVIM =3), (blaNDM and blaOXA-48=1)}, 10 isolates possessed only one gene (blaKPC =8, blaOXA-48=1 and blaVIM =1) and the remaining 26 isolates were free from these genes. Conclusion & Recommendation: In Sudan, the most common type of carbapenemase gene multidrug-resistant K. pneumoniae is KPC. Co-production of KPC, VIM, NDM and OXA-48 genes are found in K. pneumoniae. To our knowledge, this study was done for the first time in Sudan. Therefore, it is necessary to determine carbapenem resistance in K. pneumoniae isolates and take essential infection control precautions to avoid spread of this resistance.

THE ROLE OF ABERRANT METHYLATED IN GENES APC, RASSF1A AND ITGA4 FOR DIAGNOSIS OF COLORECTAL CANCER

The “gold standard” of diagnosis of colorectal cancer (CRC) is a colonoscopy. Despite the high reliability, this method is not applicable in large-scale population screening or in estimation of the disease dynamics in a particular patient. In this study, we conducted an investigation of aberrant methylation in the APC, RASSF1A and ITGA4 genes. The study included 150 pairs of tumor tissue samples with known mutation status of the RAS family genes and the surrounding histologically unchanged tissue of patients with rectal adenocarcinoma treated at the Republican Clinical Oncology Dispensary of the Ministry of Health of the Republic of Tatarstan. The methylation profiles were studied using MethyLight PCR. The most difference in the methylation between tumor and healthy tissue was observed for the ITGA4 gene (sensitivity 78%, specificity 92.7%). For the APC gene sensitivity was 32%, specificity -93.3%, for the RASSF1 gene sensitivity was 85.3%, specificity - 56.7%. Previous data on the aberrant methylation of the SEPT9 and VIM genes and new data on the APC, RASSF1A and ITGA4 genes were compared with the mutations status in the KRAS and NRAS genes. The DNA of tumor samples was significantly more often methylated in the SEPT9 (P= 0.0018) and ITGA4 (P = 0.0044) genes in the group of patients carrying mutations in the KRAS or NRAS genes, in contrast to the DNA of tumor samples of non-carriers. In statistical analysis of the effectiveness of the diagnostic test system, it was shown that our model, which includes five methylation markers (APC, RASSF1A, ITGA4, SEPT9 and VIM), has the best sensitivity and specificity (82.7% and 97.3%, respectively). The obtained model of the diagnostic test system is proposed to be used for diagnostic problems.

Detection of VIM-1 and IMP-1 genes in Klebsiella pneumoniae and relationship with biofilm formation

Klebsiella pneumoniae is an important human pathogen that is considered in recent years due to nosocomial infections resistant to treatment as well as the ability to form biofilms particularly in patients with urinary tract infection in ICU or hospital. The aim of this study was to evaluate the prevalence of VIM1, IMP1 genes and their ability to form biofilm in K. pneumoniae strains isolated from patients with urinary tract infection. In the study, using culture and biochemical methods, 1807 K. pneumoniae samples were isolated from patients with urinary tract infection hospitalized or referred to hospitals in Qom in 2013–2014. For isolation of MBL producing isolates, Double Disk Synergy Test (DDST) was used. Then MBL positive isolates were examined for the presence of VIM1, IMP1 genes using PCR method. Furthermore, all strains were investigated for biofilm formation by phenotypic microplate method. From 3165 urine samples cultured, 1807 isolates of K. pneumoniae were isolated and 109 strains (93.2%) were positive for MBL enzymes production. PCR results showed that the prevalence of VIM1 and IMP1 genes are 15.6 and 6.4%, respectively. The Phenotypic method indicated that 91.2% of isolates formed biofilm. Biofilm formation in K. pneumoniae isolates is high and there is a significant relationship between strong biofilm formation and prevalence of VIM1 and IMP1 genes. Also due to the presence of MBL genes in K. pneumoniae and horizontal transfer of genes to other bacteria, and to control the indiscriminate use of antibiotics, the hospital infection control methods must be considered.

Investigation of Methylation Changes of VIM, CXCR4, DOK7 and SPDEF Genes by Evaluation of Peripheral Blood Leukocyte DNA in Breast Cancer

Investigation of Methylation Changes of VIM, CXCR4, DOK7 and SPDEF Genes by Evaluation of Peripheral Blood Leukocyte DNA in Breast Cancer

Genetic testing for Mendelian cataract

Abstract We studied the scientific literature and disease guidelines in order to summarize the clinical utility of the genetic test for Mendelian cataract (MC). MC is caused by variations in the AGK, BFSP1, BFSP2, CHMP4B, CRYAA, CRYAB, CRYBA1, CRYBA2, CRYBA4, CRYBB1, CRYBB2, CRYBB3, CRYGC, CRYGD, CRYGS, EPHA2, EYA1, FYCO1, FOXE3, FTL, GALK1, GCNT2, GJA3, GJA8, HSF4, LEMD2, LIM2, LSS, MAF, MIP, NHS, PITX3, PAX6, SIPA1L3, SLC16A12, TDRD7, UNC45B, VIM, VSX, and WFS1 genes. The overall prevalence of congenital forms is 71 per 100 000, whereas there is insufficient data to determine the prevalence of the juvenile and age-related forms. Clinical diagnosis is based on clinical findings, age of onset, family history, ophthalmological examination and slit-lamp examination. The genetic test is useful for confirming diagnosis, and for differential diagnosis, couple risk assessment and access to clinical trials.

Triton Hodge Test for Detection of Metallo-Β-Lactamases Producing Pseudomonas Aeruginosa Isolates from Egyptian Patients with Infected Diabetic Foot

Background: One of the dreadful complications of diabetes is infected diabetic foot. Accurate detection of carbapenemase-producing Pseudomonas aeruginosa, in the form of metallo-beta-lactamases (MBLs) is necessary to determine the antibiotics for its treatment. Objectives: This study was designed to evaluate the efficacy of Triton Hodge Test (THT) versus MHT for detection of MBLs-producing P. aeruginosa isolates from infected diabetic foot patients. Methodology: Samples were collected from 500 infected diabetic footpatients. They were processed and P. aeruginosa isolates were identified. Then, carbapenem resistant P. aeruginosa (CRPA)isolates weredetected. Modified Hodge test (MHT) and Triton Hodge Test (THT) were used for detectionof MBLs. Furthermore, bla IMP , bla VIM and bla NDM MBLs genes were detected by multiplex and uniplex PCR. Resistance pattern of CRPA isolates was determined. Results: Isolation rate of P. aeruginosa from infected diabetic foot patients was 90/500 (18.00%). CRPA isolates were 16/90 (17.78%). In these isolates, the detection rate of bla NDM , bla VIM and bla IMP genes was (2/16, 12.5%), (10/16,62.5%) and (6/16,37.5%); respectively. The sensitivity, specificity and accuracy of THT in detection of NDM, VIM and IMP carbapenemases are 100 %. On the other hand, the sensitivity of MHT in detection of the studied types are 0.0%, 30.0% and 50.0%; respectively. Its specificity is 100% for all types and its accuracy is 87.5%, 56.3% and 83.1% in detection of these types. No resistance to tigecycline and colistin was detected among CRPA isolates. Conclusion: the THT is a more sensitive and accurate phenotypic test in comparison to MHT for detection of NDM, VIM and IMP carbapenemases in CRPA isolates.

The association of vimentin and fibronectin gene expression with epithelial-mesenchymal transition and tumor malignancy in colorectal carcinoma.

Colorectal cancer is the most common malignancy of the gastrointestinal tract with very high mortality. One of the most distinguishing features for the establishment of an epithelial-mesenchymal transition phenotype is the alteration of mesenchymal markers expression and structural adhesion proteins. We evaluated the significance of vimentin and fibronectin gene expression in relation to invasion and metastasis in CRC patients. Tissue specimens were collected consecutively from forty-five colorectal carcinoma patients during surgeries. Tissues were divided into two separate parts for pathological and molecular assays. In order to histological staging, tissue sections were prepared from formalin-fixed paraffin-embedded blocks and stained with Hematoxylin and Eosin. To quantify gene expression, specimens were dissected and homogenized. Moreover, SW480, SW48, SW948, Caco-2, HT-29 and LS174T as human colon cancer cell lines were obtained and cultured, then molecular analyzing was performed. As results the expression of VIM gene increased in SW480, SW48 and SW948 while it decreased in Caco-2, HT-29 and LS174T. Moreover, FN was up-regulated in Caco-2, HT-29 and SW948, while it was down-regulated in SW480, SW48 and LS174T. In tissues, vimentin and fibronectin expression significantly increased in stromal cells, whereas vimentin decreased in colonic epithelial cells and fibronectin had no significant change. Vimentin and fibronectin expression were changed in tumor tissues. It was found an association between vimentin expression with age and tumor size; over-expression in older age and decreasing in larger tumor size. Furthermore, fibronectin over-expression is correlated to older age and high tumor stages; up-regulation with increasing age and high tumor stages.

Clinical and Microbiological Characteristics of Metallo β-Lactamase Producing Pseudomonas Aueroginosa in a Sample of Egyptian Patients with Bronchiectasis

Background : Metallo-β-lactamases producing Pseudomonas aueroginosa represents a challenging concern especially in chronic colonization of bronchiectasis cases owing to progressive tissue damage, inflammation, frequent exacerbation and lack of response to antimicrobial agents. Objectives: This cross-sectional study was conducted to high-lighten the antimicrobial and molecular pattern of Pseudomonas aueroginosa isolates from bronchiectasis cases and assess their clinical outcome. Methodology: ninety-eight sputum and endotracheal aspirates were collected from bronchiectasis inpatients of Chest Department, Zagazig University Hospitals. Conventional identification and API20 NE were performed. Combined disc test and E-test were done, confirmed by PCR, to detect metallo-β-lactamases. Results : A total of 43 (43.9%) Pseudomonas were isolated, 16 (37.2%) were metallo β-lactamases and multidrug resistant. E. test had the best sensitivity and specificity. Both IMP-1 and VIM-1 genes were detected. The majority of isolates were IPM-1 positive. Conclusion : P. aeruginosa, is blamable for infection in more than 40% of bronchiectactic patients being a high-risk group. Combined disk test is a reliable, cheap and simple screening tool especially in low-resources laboratories. This study raised the concern of co-existing mechanism for both quinolone and carbapenem resistance for more investigation. Current results linked lab detection of MβL P. aeruginosa with inferior spirometry and lengthy hospital stay.

Molecular detection of β-lactamase and integron genes in clinical strains of Klebsiella pneumoniae by multiplex polymerase chain reaction.

Infections caused by β-lactamase-producing gram-negative bacteria, such as Klebsiella pneumoniae, are increasing globally with high morbidity and mortality. The aim of the current study was to determine antimicrobial susceptibility patterns and the prevalence of antibiotic resistance genes (β-lactamase and integron genes) using multiplex PCR. One-hundred K. pneumoniae isolates were collected from different clinical samples. Antibiotic susceptibility testing was performed with thirteen different antibiotics. Multiplex-PCR was used to detect β-lactamase (bla TEM, bla CTX-M, bla SHV , bla VEB, bla PER, bla GES, bla VIM, bla IMP, bla OXA, and bla KPC) and integron genes (int I, int II, and int III). The highest and lowest rate of resistance was exhibited against amikacin (93%) and imipenem (8%), respectively. The frequency of β-lactamase-positive K. pneumoniae was 37%, and the prevalence of the bla TEM, bla CTX-M, bla SHV , bla VEB, bla PER, bla GES, bla VIM, bla IMP, bla OXA, and bla KPC genes was 38%, 24%, 19%, 12%, 6%, 11%, 33%, 0%, 28%, and 23%, respectively. Of the 100 isolates, eight (8%) were positive for class I integrons; however, class II and III integrons were not detected in any of the strains. These results indicate co-carriage of a number of β-lactamase genes and antibiotic resistance integrons on the same plasmids harboring multi-drug resistance genes. It seems that these properties help to decrease treatment complications due to resistant bacterial infections by rapid detection, infection-control programs and prevention of transmission of drug resistance.

Characterization of the novel In1059 harbouring VIM gene cassette

BackgroundVIM-type enzyme encodes the most widely acquired metallo-β-lactamases in Gram- negative bacteria. To obtain current epidemiological data for integrons from enterobacteriae in hospital, the study characterizes the genetic structure in In1059 by comparison with In846 integrons harbouring VIM gene and other class 1 integrons including In37, In62, In843 and In1021 with the aim of identifying the putative mechanisms involved integron mobilization and infer evolution of relevant integrons.MethodsSix of 69 recombinant plasmids from clinical strains were found to be class 1 integrons by digestion with BamHI, drug susceptibility testing, conjugation experiments, PCR amplification, integron cloning and sequencing, genome comparison, and detection of carbapenemase activity.ResultsThe sequences of the six recombinant plasmids encoding In1021, In843, In846, In37, In62, and the novel In1059 integron had approximate lengths of ~4.8-, 4.1-, 5.1-, 5.3-, 5.3- and 6.6- kb, respectively. The genetic structures of these integrons were mapped and characterized, and the carbapenemase activities of their parental strains were assessed.ConclusionsOur results suggest that the six variable integron structures and regular variations that exist in the gene cassettes provide a putative mechanism for the integron changes. Our study has also shown that the genetic features in the integrons named above fall within a scheme involving the stepwise and parallel evolution of class 1 integron variation likely under antibiotic selection pressure in clinical settings.

The Selective Centrifugation Ensures a Better In Vitro Isolation of ASCs and Restores a Soft Tissue Regeneration In Vivo.

Autologous fat grafting procedures in plastic surgery have been extensively used to reinforce soft tissue in congenital or acquired tissue impairments. With this background, the aim of this study is firstly to examine the impact of a selective centrifugation on existing adipose stem cells (ASCs) in terms of stemness profile maintenance and, secondly, to investigate the effect of restoring volume in reconstruction on patients affected by soft tissue damage. After centrifugation, the fat graft products were separated into two layers and subsequently examined in vitro for the expression of CD34, CD90, CD117, CD105, CD29, CD31, CD44, CD73, CD133, CD14 and CD45 markers by flow cytometry and gene expression analyses were performed for Sox2, WNT3A, END, CD44, FUT4, COLL1, CTNNB1, hbEGF, KRTLG, MMP2 and VIM genes. The results showed that in the middle-high density (MHD) layer there was a peak concentration of ASCs, compared to another layer obtained after centrifugation. Research carried out on patients under treatment for soft tissue regeneration using cells obtained from MHD layer selection will be fundamental in comparative analysis. These studies will lead to an adequate standardization of outcomes, provided that treatment is performed through cell selection. Therefore, a unique procedure in tissue reconstruction and regeneration through fat grafting is presented here.

Environmental pollution with antimicrobial agents from bulk drug manufacturing industries in Hyderabad, South India, is associated with dissemination of extended-spectrum beta-lactamase and carbapenemase-producing pathogens.

High antibiotic and antifungal concentrations in wastewater from anti-infective drug production may exert selection pressure for multidrug-resistant (MDR) pathogens. We investigated the environmental presence of active pharmaceutical ingredients and their association with MDR Gram-negative bacteria in Hyderabad, South India, a major production area for the global bulk drug market. From Nov 19 to 28, 2016, water samples were collected from the direct environment of bulk drug manufacturing facilities, the vicinity of two sewage treatment plants, the Musi River, and habitats in Hyderabad and nearby villages. Samples were analyzed for 25 anti-infective pharmaceuticals with liquid chromatography-tandem mass spectrometry and for MDR Gram-negative bacteria using chromogenic culture media. In addition, specimens were screened with PCR for bla VIM, bla KPC, bla NDM, bla IMP-1, and bla OXA-48 resistance genes. All environmental specimens from 28 different sampling sites were contaminated with antimicrobials. High concentrations of moxifloxacin, voriconazole, and fluconazole (up to 694.1, 2500, and 236,950µg/L, respectively) as well as increased concentrations of eight other antibiotics were found in sewers in the Patancheru-Bollaram industrial area. Corresponding microbiological analyses revealed an extensive presence of extended-spectrum beta-lactamase and carbapenemase-producing Enterobacteriaceae and non-fermenters (carrying mainly bla OXA-48, bla NDM, and bla KPC) in more than 95% of the samples. Insufficient wastewater management by bulk drug manufacturing facilities leads to unprecedented contamination of water resources with antimicrobial pharmaceuticals, which seems to be associated with the selection and dissemination of carbapenemase-producing pathogens. The development and global spread of antimicrobial resistance present a major challenge for pharmaceutical producers and regulatory agencies.

A study on antibiotics resistance and carbapenemase genotype of Acinetobacter baumannii in intensive care unit of Qingdao

Objective To investigate antibiotics resistant characteristics and carbapenemases genotype of Acinetobacter baumannii in Intensive Care Unit (ICU), so as to provide theoretical basis for clinical prevention and treatment. Methods Retrospective study was made on 90 non-duplicated clinical isolates of Acinetobacter baumannii, which were collected From January 2013 to January 2014 in three tertiary hospitals of Qingdao. All strains were identified by VITEK2 automated microbiology analyzer; K-B method was used to do drug susceptibility test; polymerase chain reaction (PCR) was used to amplify the OXA-23, OXA-24, OXA-51, OXA-58, KPC-2, VIM, IMP genes, and the positive products of genes were sequenced; the chi-square test was used to compare the difference of the resistance rates. Results The detection rate of multi-drug resistant A. baumannii (MDRAB)and Pan-drug resistant A. baumannii (PDRAB)was 61.11% (55/90) and 17.78% (16/90). In the 32 strains of imipenem-resistant Acinetobacter baumannii, the resistant rates to Cefoperazone/sulbactam, Polymyxin B was lower, while the resistant rates to other drugs tested were more than 85%. The difference of the resistance rates to 9 drugs between imipenem resistant group and Imipenem sensitive group were statistically significant (P≤0.05). PCR result showed: 32 strains detected OXA-51 gene, 28 strains detected OXA-23 gene, and 3 strains detected VIM gene, the detection rates of which were 100%, 87.50% and 9.38% respectively.All strains were not detected OXA-24, OXA-58, KPC-2 and IMP genes. The sequenced results were absolutely homology with the corresponding genes in genbank. Conclusions The resistance of A. baumannii in ICU is serious in this region, especially imipenem-resistant A. baumannii, which were nearly no-sensitive to most of the drugs commonly used in clinical. The gene existence of carbapenemase and carbapenemase producing is one of the main resistance mechanism of Acinetobacter baumannii to carbapenem antibiotics. OXA-23 was the major genotypes in this region.(Chin J Lab Med, 2017, 40: 216-220) Key words: Acinetobacter baumannii; Drug resistance, bacterial; Bacterial proteins; Beta-Lactamases; Genotype; Intensive care units

TMPRSS2:ERG gene fusion variants induce TGF-β signaling and epithelial to mesenchymal transition in human prostate cancer cells.

TMPRSS2:ERG (T/E) gene fusions are present in approximately 50% of all prostate cancer (PCa) cases. The expression of fusion mRNAs from distinct T/E variants is associated with clinicopathological parameters, while the underlying molecular processes remain unclear. We characterized the molecular mechanisms and functional implications caused by doxycycline (Dox)-inducible overexpression of the frequent T/E III and VI fusion variants in LNCaP cells. Induction of T/E expression resulted in increased cellular migratory and invasive potential, and reduced proliferation and accumulation in G1 phase. T/E overexpressing cells showed epithelial-to-mesenchymal transition (EMT), as demonstrated by upregulation of TGF-β and WNT pathway genes, mesenchymal markers, and increased phosphorylation of the p38 MAPK. Augmented secretion of TGF-β1 and –β2, and T/E-mediated regulation of ALK1, a member of the TGF-β receptor family, was detected. ALK1 inhibition in T/E overexpressing cells blocked p38 phosphorylation and reduced the expression of the TGF-β target genes VIM, MMP1, CDH2, and SNAI2. We found a T/E variant VI-specific induction of miR-503 associated with reduced expression of SMAD7 and CDH1. Overexpression of miR-503 led to increased levels of VIM and MMP1. Our findings indicate that TGF-β signaling is a major determinant of EMT in T/E overexpressing LNCaP cells. We provide evidence that T/E VI-specific transcriptional modulation by miR-503 accounts for differences in the activation of EMT pathway genes, promoting the aggressive phenotype of tumors expressing T/E variant VI. We suggest that ALK1-mediated TGF-β signaling is a novel oncogenic mechanism in T/E positive PCa.

Prevalence, antibiotic susceptibility and characterization of antibiotic resistant genes among carbapenem-resistant Gram-negative bacilli and yeast in intestinal flora of cancer patients in North Lebanon.

The emergence and spread of carbapenem-resistant bacteria are a significant clinical and public health concern. The aim of the study is to determine the prevalence of intestinal carriage of carbapenem-resistant bacteria and yeasts in cancer patients under chemotherapy. 41 stool samples collected from cancer patients in Nini hospital in Tripoli, North Lebanon have been analyzed. After isolating yeasts and carbapenem-resistant bacteria, a biochemical identification and antimicrobial susceptibility profile were determined. The mechanism of enzymatic carbapenem-resistance was detected by searching for carbapenemases by both Hodge test and PCR assays. The association of several mechanisms of resistance was also searched. 46.3% (19/41) of patients were colonized by yeast. Candida glabrata (6/19) was the major species. The prevalence of carbapenem-resistant bacteria was 24.4% (10/41) including Escherichia coli (5/10), Enterobacter cloacae (1/10), Enterobacter aerogenes (1/10) Edwardsiella hoshinae (1/10) Pantoea agglomerans (1/10) and Pseudomonas stutzeri (1/10). PCR and sequencing of the amplified fragments revealed that Pseudomonas stutzeri (1/1) carried VIM gene and Enterobacter aerogenes (1/1) and E. coli (1/5) carried OXA-48 gene. The other Enterobacteriaceae were resistant to carbapenems by mechanisms other than a carbapenemase including hyperproduction of cephalosporinase (4/10), extended spectrum beta-lactamases (1/10) and both cephalosporinase and extended spectrum beta-lactamases (2/10). High prevalence of intestinal carriage of carbapenem-resistant bacteria and yeasts were detected in cancer patients under chemotherapy. In order to prevent the development of endogenous infection and the dissemination of antimicrobial resistance, an implementation of antibiotic stewardship programs and infection control measures is required in hospitals particularly in the department of chemotherapy.

Carbapenemase-producing Klebsiella pneumoniae bloodstream infection in critically ill patients: risk factors and predictors of mortality.

A significant increase in carbapenemase-producing Klebsiella pneumoniae (CP-Kp) bacteraemias has been observed worldwide. The objective of the present work was to study the risk factors and predictors of mortality of CP-Kp bacteraemias among critically ill patients. During a 4-year period (2012-3015), a matched 1:2 case-control study was conducted. Klebsiella pneumoniae was identified by Vitek 2 technology. Antibiotic susceptibility was performed by the agar disc diffusion method and Etest. The presence of the bla KPC, bla VIM and bla NDM genes was confirmed by polymerase chain reaction (PCR). Epidemiologic data were collected from the intensive care unit (ICU) computerised database. One hundred and thirty-nine patients who developed a CP-Kp bacteraemia were matched with 278 patients. The majority of isolates (128; 92.1%) carried the bla KPC gene, seven carried both bla KPC and bla VIM, three bla VIM and one carried bla NDM. Risk factors for the development of CP-Kp bacteraemia were administration of tigecycline and number of antibiotics administered prior to CP-Kp bacteraemia. Overall, the 30-day mortality was 36.0%. Multivariate analysis revealed septic shock, Simplified Acute Physiology Score II (SAPS II) upon infection onset, adjunctive corticosteroid administration and parenteral nutrition as independent predictors of mortality, while treatment with a combination of appropriate antibiotics was identified as a predictor of good prognosis. Among septic shock patients (n = 74), Sequential Organ Failure Assessment (SOFA) score upon infection onset, adjunctive corticosteroid administration and strain carrying the bla KPC gene were independently associated with mortality, while the administration of combination treatment was identified as a predictor of a good prognosis. The administration of tigecycline predisposes to the induction of bacteraemia. Appropriate antibiotic treatment is associated with better survival, while concomitant corticosteroid treatment is associated with mortality.

Emergence of Imipenem-ResistantPseudomonas aeruginosaClinical Isolates from Egypt CoharboringVIMandIMPCarbapenemases

Pseudomonas aeruginosa is an important human pathogen and the leading cause of nosocomial infections. P. aeruginosa is characterized by massive intrinsic resistance to a multiple classes of antibiotics with carbapenems being the most potent inhibitor of P. aeruginosa and considered the first choice for its treatment. Therefore, it is crucial to investigate novel mechanisms of resistance of P. aeruginosa to carbapenems for achieving successful therapy. A total of 114 P. aeruginosa isolates from two university hospitals in Egypt were recruited in this study. Antimicrobial susceptibility testing revealed that 50 isolates (43.8%) exhibited multidrug-resistant (MDR) phenotype, of them 14 isolates (12.2%) were imipenem (IPM)-resistant. Of these 14 isolates, 13 isolates (11.4%) exhibited the metallo-β-lactamase (MBL) phenotype. MBLs encoding genes, VIM and IMP, were identified by PCR. PCR results revealed that four isolates harbored the VIM gene alone, one isolate harbored IMP gene alone, and four isolates harbored both genes. The correct size of PCR products of VIM and IMP genes (390 and 188 bp, respectively) were sequenced to confirm results of PCR and to look for any possible polymorphism among MBL genes of tested isolates. Data analysis of these sequences showed 100% identity of nucleotide sequences of MBL genes among tested Egyptian patients. To our knowledge, this is the first report of IMP carbapenemase-encoding gene in Africa and the first detection of the emergence of P. aeruginosa coproducing VIM and IMP genes in Egypt.

The Prevalence of Extended-Spectrum β-lactamase (ESBL) and Carbapenem-Resistant Enterobacteriaceae (CRE) Isolates in Positive Blood Cultures of Patients in A Teaching Hospital in Kuwait Over A 2-year Period

Extended spectrum β-lactamase (ESBL)-producing organisms and carbapenem resistant Enterobacteriaceae (CRE) pose unique challenges to clinical microbiologists, infectious diseases physicians, infection control professionals and antibacterial-discovery scientists. This study was undertaken to determine the prevalence of extended-spectrum β-lactamase (ESBL)-producing and carbapenemase-positive members of the family Enterobacteriaceae isolated from positive blood cultures of patients in our hospital. Enterobacteriaceae isolates from proven cases of blood stream infections (BSI) from January 2014 to December 2015 were included in the study. They were identified and tested for their susceptibilities to 18 antibiotics by conventional methods and semi-automated VITEK-2 system. ESBL and carbapenem resistance (CR) were detected by phenotypic and molecular PCR methods, and characterized by sequencing. A total of 4133 blood cultures were positive. Of these, 543 (13.1%) were positive for Enterobacteriaceae, 135 (24.9%) of which were ESBL producers and 28 (5.2%) identified as CRE. The most common of the ESBL-producing isolates were Escherichia coli (81:60%) and Klebsiella pneumoniae (44:32.6%). Resistance rates of E. coli to the cephalosporins ranged from 47–100%, ciprofloxacin 70.4%, imipenem 2.5% and meropenem 1.2%. Higher proportions of K. pneumoniae were resistant to the cephalosporins (68.2–100%), imipenem (27.3%) and meropenem (22.7%). Sequence analysis of the ESBL revealed CTX-M-15 as the commonest enzyme (75%). The CRE harbored mainly the VIM (28.6%) and NDM-1 (35.7%) genes. Our study demonstrates a high prevalence of ESBL-producing Enterobacteriaceae isolates of the CTX-M-15 variety and equally high prevalence of CRE, predominantly VIM and NDM-1 producers, among blood culture isolates at our hospital. All authors: No reported disclosures.