Increased Levels Of Resistance Research Articles

Shifting trends in bacteriology and antimicrobial resistance among gastrointestinal fistula patients in China: an eight-year review in a tertiary-care hospital

BackgroundThe purpose of this study was to determine the shifting trends in bacteriology and antimicrobial resistance of infectious specimens isolated from gastrointestinal (GI) fistula patients over eight years in China.MethodsWe retrospectively reviewed the microbial records of intra-abdominal specimens at a teaching hospital from 2008 to 2015. Study period was divided into the first half (2008–2011) and the second half (2012–2015). All isolates underwent antibiotic susceptibility testing by the micro dilution method.ResultsA total of 874 intra-abdominal isolates were consecutively collected from 502 GI fistula patients (mean age, 46.5 years, 71.1% male) during the study period. Patients in the second study period (2012–2015) were older (>65 years) and more likely to have experienced cancer. Over the entire study period, most infections were caused by E. coli (24.2%) and K. pneumonia (14.1%). There was a significant decrease in the proportion E. coli isolates that were extended- spectrum beta-lactamase (ESBL)-positive (P = 0.026). The proportion of E. coli resistant to imipenem increased from 14.3% in 2008–2011 to 25.9% in 2012–2015 (P = 0.037). Imipenem resistance prevalence was higher in ESBL-negative bacteria than ESBL-positive bacteria for both E. coli and K. pneumonia (P < 0.001). In Enterococcus, significant increase in resistance to ampicillin (P = 0.01) and moxifloxacin (P = 0.02) over time were observed. In Staphylococcus and fungi, rates of antibiotic resistance did not significantly change over the study period.ConclusionsGram-negative bacteria predominated as causative agents of intra-abdominal infections in GI fistula patients, and there was an increase in levels of resistance to certain antibiotics, particularly carbapenems. Infection control and source control are important tools available to surgeons to prevent the emergence of antibiotic-resistant pathogens.

Pseudomonas aeruginosa prevalence, antibiotic resistance and antimicrobial use in Chinese burn wards from 2007 to 2014.

ObjectiveTo assess the application of antibacterial agents, alongside pathogen prevalence and Pseudomonas aeruginosa drug resistance, with the aim of understanding the impact of inappropriate antibacterial use.MethodsThis retrospective study assessed bacteria from wounds, catheters, blood, faeces, urine and sputum of hospitalized patients in burn wards between 2007 and 2014. The intensity of use of antibacterial agents and resistance of P. aeruginosa to common anti-Gram-negative antibiotics were measured.ResultsAnnual detection rates of Staphylococcus aureus were significantly decreased, whereas annual detection rates of P. aeruginosa and Klebsiella pneumoniae were significantly increased. Multidrug-resistant strains of P. aeruginosa were increased. The intensity of use of some anti-Gramnegative antibiotics positively correlated with resistance rates of P. aeruginosa to similar antimicrobials.ConclusionIn burn wards, more attention should be paid to P. aeruginosa and K. pneumoniae. The use of ciprofloxacin, ceftazidime and cefoperazone/sulbactam should be limited to counter the related increase in resistance levels.

Predicting antibiotic prescription after symptomatic treatment for urinary tract infection: development of a model using data from an RCT in general practice

BackgroundUncomplicated urinary tract infection (UTI) is often treated with antibiotics, resulting in increasing resistance levels. A randomised controlled trial showed that two-thirds of females with UTI treated symptomatically recovered without subsequent antibiotic treatment.AimTo investigate whether there are differences between females with a UTI who were subsequently prescribed antibiotics and those who recovered with symptomatic treatment only, and to develop a model to predict those who can safely and effectively be treated symptomatically.Design and settingThis is a subgroup analysis of females assigned to ibuprofen in a UTI trial in general practices.MethodMultiple logistic regression analysis was used to select variables for a prediction model, The discriminative value of the model was estimated by the area under the receiver operator curve (AUC) and the effects of different thresholds were calculated within the model predicting antibiotic prescription and need for follow-up visits.ResultsOf the 235 females in the ibuprofen group, 79 were subsequently prescribed antibiotics within 28 days of follow-up. The final model included five predictors: urgency/frequency, impaired daily activities, and positive dipstick test results for erythrocytes, leucocytes, and nitrite. The AUC was 0.73 (95% CI = 0.67 to 0.80). A reasonable threshold for antibiotic initiation would result in 58% of females presenting with UTI being treated with antibiotics. Of the remaining females, only 6% would return to the practice because of symptomatic treatment failure.ConclusionThe present model revealed moderately good accuracy and could be the basis for a decision aid for GPs and females to find the treatment option that fits best.

Identification of genes differentially expressed during early interactions between the stem rot fungus (Sclerotium rolfsii) and peanut (Arachis hypogaea) cultivars with increasing disease resistance levels

Sclerotium rolfsii, a destructive soil-borne fungal pathogen causes stem rot of the cultivated peanut, Arachis hypogaea. This study aimed to identify differentially expressed genes associated with peanut resistance and fungal virulence. Four peanut cultivars (A100-32, Georgia Green, GA-07W and York) with increasing resistance levels were inoculated with a virulent S. rolfsii strain to study the early plant–pathogen interaction. 454 sequencing was performed on RNAs from infected tissue collected at 4 days post inoculation, generating 225,793 high-quality reads. Normalized read counts and fold changes were calculated and statistical analysis used to identify differentially expressed genes. Several genes identified as differential in the RNA-seq experiment were selected based on functions of interest and real-time PCR employed to corroborate their differential expression. Expanding the analysis to include all four cultivars revealed a small but interesting set of genes showing colinearity between cultivar resistance and expression levels. This study identified a set of genes possibly related to pathogen response that may be useful marker assisted selection or transgenic disease control strategies. Additionally, a set of differentially expressed genes that have not been functionally characterized in peanut or other plants and warrant additional investigation were identified.

Genotypic and phenotypic analyses of a Pseudomonas aeruginosa chronic bronchiectasis isolate reveal differences from cystic fibrosis and laboratory strains.

BackgroundPseudomonas aeruginosa is an environmentally ubiquitous Gram-negative bacterium and important opportunistic human pathogen, causing severe chronic respiratory infections in patients with underlying conditions such as cystic fibrosis (CF) or bronchiectasis. In order to identify mechanisms responsible for adaptation during bronchiectasis infections, a bronchiectasis isolate, PAHM4, was phenotypically and genotypically characterized.ResultsThis strain displays phenotypes that have been associated with chronic respiratory infections in CF including alginate over-production, rough lipopolysaccharide, quorum-sensing deficiency, loss of motility, decreased protease secretion, and hypermutation. Hypermutation is a key adaptation of this bacterium during the course of chronic respiratory infections and analysis indicates that PAHM4 encodes a mutated mutS gene responsible for a ~1,000-fold increase in mutation rate compared to wild-type laboratory strain P. aeruginosa PAO1. Antibiotic resistance profiles and sequence data indicate that this strain acquired numerous mutations associated with increased resistance levels to β-lactams, aminoglycosides, and fluoroquinolones when compared to PAO1. Sequencing of PAHM4 revealed a 6.38 Mbp genome, 5.9 % of which were unrecognized in previously reported P. aeruginosa genome sequences. Transcriptome analysis suggests a general down-regulation of virulence factors, while metabolism of amino acids and lipids is up-regulated when compared to PAO1 and metabolic modeling identified further potential differences between PAO1 and PAHM4.ConclusionsThis work provides insights into the potential differential adaptation of this bacterium to the lung of patients with bronchiectasis compared to other clinical settings such as cystic fibrosis, findings that should aid the development of disease-appropriate treatment strategies for P. aeruginosa infections.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-2069-0) contains supplementary material, which is available to authorized users.

Rise of multiple insecticide resistance in Anopheles funestus in Malawi: a major concern for malaria vector control.

BackgroundDeciphering the dynamics and evolution of insecticide resistance in malaria vectors is crucial for successful vector control. This study reports an increase of resistance intensity and a rise of multiple insecticide resistance in Anopheles funestus in Malawi leading to reduced bed net efficacy.MethodsAnopheles funestus group mosquitoes were collected in southern Malawi and the species composition, Plasmodium infection rate, susceptibility to insecticides and molecular bases of the resistance were analysed.ResultsMosquito collection revealed a predominance of An. funestus group mosquitoes with a high hybrid rate (12.2 %) suggesting extensive species hybridization. An. funestus sensu stricto was the main Plasmodium vector (4.8 % infection). Consistently high levels of resistance to pyrethroid and carbamate insecticides were recorded and had increased between 2009 and 2014. Furthermore, the 2014 collection exhibited multiple insecticide resistance, notably to DDT, contrary to 2009. Increased pyrethroid resistance correlates with reduced efficacy of bed nets (<5 % mortality by Olyset® net), which can compromise control efforts. This change in resistance dynamics is mirrored by prevalent resistance mechanisms, firstly with increased over-expression of key pyrethroid resistance genes (CYP6Pa/b and CYP6M7) in 2014 and secondly, detection of the A296S-RDL dieldrin resistance mutation for the first time. However, the L119F-GSTe2 and kdr mutations were absent.ConclusionsSuch increased resistance levels and rise of multiple resistance highlight the need to rapidly implement resistance management strategies to preserve the effectiveness of existing insecticide-based control interventions.Electronic supplementary materialThe online version of this article (doi:10.1186/s12936-015-0877-y) contains supplementary material, which is available to authorized users.

Emergence of sulfadoxine–pyrimethamine resistance in Indian isolates of Plasmodium falciparum in the last two decades

Genotyping the sulfadoxine–pyrimethamine (SP) genes will help in identifying the genes under drug selection and the emergence of resistance in dhfr and dhps genes. India is an important hotspot for studying malaria due to the immense climatic diversity prevalent in the country. The central and eastern parts of the country are most vulnerable sites where malaria cases are reported throughout the year. From different regions of the country 173 field isolates were genotyped at various loci in dhfr and dhps genes collected between 1994 and 2013. This encompasses the period before antimalarial resistance emerged and the period after the use of combination therapy was made mandatory in the country. We observed the rise of resistant SP alleles from very low frequencies (in the year 1994) to steadily rising (in the year 2000) and maintaining this increasing trend subsequently (in the year 2013) as shown by the sequence analysis of dhfr and dhps genes. This study assessed the prevalence of mutations in dhfr and dhps genes associated with SP resistance in samples indicative of increase in resistance levels of Plasmodium falciparum to SP even after the change in malaria treatment policy in the country.

The challenge of efflux-mediated antibiotic resistance in Gram-negative bacteria.

The global emergence of multidrug-resistant Gram-negative bacteria is a growing threat to antibiotic therapy. The chromosomally encoded drug efflux mechanisms that are ubiquitous in these bacteria greatly contribute to antibiotic resistance and present a major challenge for antibiotic development. Multidrug pumps, particularly those represented by the clinically relevant AcrAB-TolC and Mex pumps of the resistance-nodulation-division (RND) superfamily, not only mediate intrinsic and acquired multidrug resistance (MDR) but also are involved in other functions, including the bacterial stress response and pathogenicity. Additionally, efflux pumps interact synergistically with other resistance mechanisms (e.g., with the outer membrane permeability barrier) to increase resistance levels. Since the discovery of RND pumps in the early 1990s, remarkable scientific and technological advances have allowed for an in-depth understanding of the structural and biochemical basis, substrate profiles, molecular regulation, and inhibition of MDR pumps. However, the development of clinically useful efflux pump inhibitors and/or new antibiotics that can bypass pump effects continues to be a challenge. Plasmid-borne efflux pump genes (including those for RND pumps) have increasingly been identified. This article highlights the recent progress obtained for organisms of clinical significance, together with methodological considerations for the characterization of MDR pumps.

Prognostic significance of novel katG mutations in Mycobacterium tuberculosis

BackgroundBy using whole genome sequencing (WGS), researchers are beginning to understand the genetic diversity of Mycobacterium tuberculosis (MTB) and its consequences for the diagnosis of multidrug-resistant tuberculosis (MDR–TB) on a genomic scale. The Global Consortium for Drug-resistant TB Diagnostics (GCDD) conducted a genome scale variant analyses of 366 clinical MTB genomes (mostly MDR/XDR [extensively drug resistant]) from four countries in order to inform the development of rapid molecular diagnostics. This project has been extended by performing an evolutionary analysis of isoniazid (INH)-resistant isolates for prognostic purposes. Methods151 (130 INHR, 21 INHS) clinical MTB isolates from India (19: 17 INHR, 2 INHS), Moldova (48: 42 INHR, 6 INHS), the Philippines (26: 20 INHR, 6 INHS), and South Africa (58: 51 INHR, 7 INHS) were included in this study. INH drug susceptibility was determined by using MGIT 960 and WHO (World Health Organization)-recommended critical concentration of 0.1mg/L. Isolates were sequenced using PacBio RS WGS platform. A genome-wide variant analysis was conducted using a proprietary pipeline (PacDAP) developed at San Diego State University. To infer the amino acid changes in katG that confer resistance, PAML was utilized to detect sites in silico that are under positive selection. The dN/dS method was used in combination with Bayes empirical Bayes to determine sites under positive selection and Chi-Squared analysis to determine the significance of the selected sites. ResultsPacDAP variant analysis revealed 22 novel catalase-peroxidase (katG product) mutations. Of these, 14 were single nucleotide polymorphisms, while 8 novel mutations appeared in combination with katG S315T and/or with inhA promoter C-15T. These SNPs have not been previously reported. Additionally, 11 previously observed, but uncommon, katG mutations were also observed in these clinical isolates. These results suggest that 17 amino acids in the enzyme are under positive selective pressure; most significantly in South Africa and the Philippines. No selective pressure on codons other than 315 was observed in isolates from Moldova. Due to the low number of isolates from India, the significance of the sites under positive selection was low and no prediction for India could be made based on this study. ConclusionsEleven of the 14 SNPs are resistance conferring, and it is believed that the remaining 8 combinatorial mutations are either compensatory in nature or, in combination with known SNPs, could increase resistance levels. Positive selection results indicate a diversifying evolutionary path to resistance more in line with long tail statistics and therefore indicate a departure from the traditional point mutation (or “hotspot”) model that current molecular diagnostics are based on. Positive selection pressures indicate a future with elevated diagnostic and prognostic significance of the “long tail” (i.e., alternative mechanisms of resistance) and potentially diminishing significance of the canonical mutations (especially in South Africa and the Philippines), which could have significant future implications on narrowly targeting molecular diagnostics.

Effects of stacked quantitative resistances to downy mildew in lettuce do not simply add up.

In a stacking study of eight resistance QTLs in lettuce against downy mildew, only three out of ten double combinations showed an increased resistance effect under field conditions. Complete race nonspecific resistance to lettuce downy mildew, as observed for the nonhost wild lettuce species Lactuca saligna, is desired in lettuce cultivation. Genetic dissection of L. saligna's complete resistance has revealed several quantitative loci (QTL) for resistance with field infection reductions of 30-50 %. To test the effect of stacking these QTL, we analyzed interactions between homozygous L. saligna CGN05271 chromosome segments introgressed into the genetic background of L. sativa cv. Olof. Eight different backcross inbred lines (BILs) with single introgressions of 30-70 cM and selected predominately for quantitative resistance in field situations were intercrossed. Ten developed homozygous lines with stacked introgression segments (double combinations) were evaluated for resistance in the field. Seven double combinations showed a similar infection as the individual most resistant parental BIL, revealing epistatic interactions with 'less-than-additive' effects. Three double combinations showed an increased resistance level compared to their parental BILs and their interactions were additive, 'less-than-additive' epistatic and 'more-than-additive' epistatic, respectively. The additive interaction reduced field infection by 73 %. The double combination with a 'more-than-additive' epistatic effect, derived from a combination between a susceptible and a resistant BIL with 0 and 30 % infection reduction, respectively, showed an average field infection reduction of 52 %. For the latter line, an attempt to genetically dissect its underlying epistatic loci by substitution mapping did not result in smaller mapping intervals as none of the 22 substitution lines reached a similar high resistance level. Implications for breeding and the inheritance of L. saligna's complete resistance are discussed.

Extended-spectrum β-lactamase and carbapenemase-producing Aeromonas species in wild animals from Portugal

Aeromonas are Gram-negative, facultative-anaerobic, non-spore-forming, glucose-fermenting, oxidase- and catalase-positive rods (Martin-Carnahan and Joseph 2005). Apart from fish, which are widely reported hosts for Aeromonads, insects, crustaceans, reptiles, birds and mammals...

Resistance to selected beta-lactam antibiotics

Susceptibility in vitro and trends in resistance to antimicrobials were determined by a dilution micromethod in a group of Actinobacillus pleuropneumoniae, Pasteurella multocida, Mannheimia haemolytica and Escherichia coli isolates from clinical cases of cattle and swine diseases in the Czech Republic from 2007 to 2011. A high susceptibility of pig and cattle respiratory pathogens to antimicrobials was found, with the exception of the moderate prevalence of M. haemolytica resistance to ampicillin. In contrast to respiratory pathogens, low susceptibility of E. coli of pig and cattle isolates to ampicillin and amoxicillin/clavulanic acid was noted. Regarding resistance trends, an increase in levels of resistance among E. coli isolates to ampicillin and amoxicillin/clavulanic acid was identified, but the resistance of respiratory isolates was low, with the exception of M. haemolytica.For the period of 2007–2011, there was a significant and almost continuous increase in sales (compared with population correction unit) of ceftiofur, cefquinome and other beta lactams for pigs. Consumption peaked in 2010. In the case of amoxicillin in combination with clavulanic acid, data showed a significant decrease in sales from 2007 to 2008, followed by a period of fluctuation. In cattle, within the groups of 3rd and 4th generation cephalosporins and for the whole group of other betalactams for the period of 2007–2011, there was a significant and almost continuous increase in sales (compared with population correction unit). Consumption peaked in 2010. In the case of ceftiofur, there was a huge increase noted from 2010. In the case of amoxicillin in combination with betalactamase inhibitor (clavulanic acid) data shows a significant decrease from 2007 to 2008, followed by a period of fluctuation in sales.

Molecular analysis of ciprofloxacin resistance among non-typhoidal Salmonella with reduced susceptibility to ciprofloxacin isolated from patients at a tertiary care hospital in Kuala Lumpur, Malaysia.

We investigated the prevalence of non-typhoidal Salmonella (NTS) with "reduced susceptibility to ciprofloxacin" (RS-Cip) (minimum inhibitory concentration [MIC], 0.12-1.0 μg/mL) as well as their resistance genes in 75 NTS isolates (53 from stool, 21 from blood, and 1 from urine) from patients at a tertiary care Malaysian hospital between January and December 2009. RS-Cip was detected in 24/75 (32.0%) isolates. Using the ciprofloxacin MIC interpretive criteria for Salmonella in the Clinical and Laboratory Standards Institute 2013 guidelines, 51/75 (68.0%) isolates were found to be sensitive, 22/75 (29.3%) were intermediate, and 2/75 (2.7%) were resistant to ciprofloxacin. The 24 isolates that were intermediate or resistant to ciprofloxacin were the same isolates categorized as having RS-Cip. Among the 23 tested isolates with RS-Cip, the qnrS gene was detected in 17/23 (73.9%) and single gyrA mutations were detected in 6/23 (26.1%) (Asp87Tyr [n = 3], Asp87Asn [n = 2], and Ser83Phe [n = 1]). A parC (Thr57Ser) mutation was detected in 13/23 (56.5%) isolates, coexisting with either a qnrS gene or a gyrA mutation. The high incidence of the qnrS gene among isolates with RS-Cip needs to be monitored because qnr genes can spread via plasmids and aid in the emergence of increased resistance levels.

Interactions between copy number and expression level of genes involved in fluconazole resistance in Candida glabrata

Objectives: This study aimed to elucidate the relative involvement of drug resistance gene copy number and overexpression in fluconazole resistance in clinical C. glabrata isolates using a population-based approach.Methods: Fluconazole resistance levels were quantified using the minimal inhibitory concentration (MIC) via Etest method. Both gene expression levels and gene copy number of CgCDR1, CgPDH1, CgERG11, and CgSNQ2 were assessed via quantitative real-time PCR. The influence of the main effects and first-level interactions of both the expression level and copy number of these genes on fluconazole resistance levels were analyzed using a multivariate statistical model.Results: Forty-three C. glabrata isolates were collected from 30 patients during in a hospital survey. In the multivariate analysis, C. glabrata fluconazole MICs were independently increased by CgSNQ2 overexpression (p < 10−4) and the interaction between CgPDH1 gene copy number and CgPDH1 expression level (p = 0.038). In contrast, both CgPDH1 overexpression (p = 0.049) and the interaction between CgSNQ2 and CgERG11 expression (p = 0.003) led to a significant decrease in fluconazole MICs.Conclusion: Fluconazole resistance in C. glabrata involves complex interactions between drug resistance gene expression and/or copy number. The population-based multivariate analysis highlighted the involvement of the CgSNQ2 gene in fluconazole resistance and the complex effect of the other genes such as PDH1 for which overexpression was associated with reduced fluconazole resistance levels, while the interaction between PDH1 overexpression and copy number was associated with increased resistance levels.

Antimicrobial Resistance Patterns of Bacterial Isolates in a Medical College Hospital of Bangladesh, 2005-2006

Current knowledge on antimicrobial susceptibility pattern is essential for quick and appropriate therapy. But, despite the seriousness of the problem, available data on antibiotic resistant pathogens are limited in Bangladesh. For this reason, the present study have been undertaken to determine the antibiotic susceptibility patterns of isolated bacterial strains to guide the clinician in selecting the best antimicrobial agent for an individual patient and to accumulate epidemiological information for public health importance. Isolates were identified by conventional methods from different samples of urine, blood, pus, wound swab, stool, high vaginal swab and pleural fluid from patients attending to Sir Salimullah Medical College and Hospital in Dhaka, Bangladesh between January 2005 and October 2006. Antimicrobials susceptibility testing was performed by Kirby Bauer’s disc diffusion method and reported according to CLSI guidelines. Of the 1592 different tested samples, 779 showed growth of pathogens, among which the most prevalent gram negative bacilli were Escherichia coli (71%) followed by Pseudomonas (8%) and Klebsiella (5%). Staphylococcus aureus (11%) was found as the most important gram positive resistant pathogens. The resistance pattern among gram negative bacilli was observed high against ampicillin, cephalexin, cotrimoxazole, nalidixic acid and ciprofloxacin. Staphylococcus showed almost same trend except ciprofloxacin and nalidixic acid. Most of the isolates were resistant to 4 or more number of antibiotics. Available antimicrobials with good activity against resistant pathogens include ceftriaxone and imipenem. The role of fluoroquinolones in the empiric treatment of bacterial infections is also being limited by new resistance patterns and increasing resistance levels, resulting a considerable economic and health burdens on our country.DOI: http://dx.doi.org/10.3329/bjmm.v7i1.19316 Bangladesh J Med Microbiol 2013; 07(01): 15-19

A QTL on chromosome 2DS of ‘Sumai 3’ increases susceptibility to Fusarium head blight in wheat

Much effort has been invested in identifying molecular markers in wheat (Triticum aestivum L.) linked to quantitative trait loci (QTL) that confer resistance to Fusarium head blight (FHB) caused by Fusarium graminearum Schwabe [teleomorph Gibberella zeae (Schwein) Petch]. Even after several generations of crossing and selection by many wheat breeding programs, resistance of the Chinese spring wheat cultivar ‘Sumai 3’ (PI 481542) remains among the most effective. It therefore seems that undocumented resistance QTL present in Sumai 3 were not detected in various mapping studies. Using an extremely susceptible Tibetan landrace (‘Y1193-6’; unknown pedigree) in the creation of a mapping population with Sumai 3, the objective of this research was to identify undocumented resistance QTL in Sumai 3. This was accomplished through collecting disease index (DI) and Fusarium damaged kernel (FDK) phenotypic values along with 305 Diversity Array Technology (DArT) and 52 Simple Sequence Repeat (SSR) marker genotypes on 160 F2:6 recombinant inbred lines (RILs). Disease response evaluations were based on four (two greenhouse and two field) experiments where spray inoculation methods were used. Three QTL were identified on chromosome arms 3BS, 6BL and 2DS explaining 26.1, 10.7 and 18.9% of the phenotypic variation for DI, respectively. The same QTL were also significantly associated with reduced FDK scores and explained 28.0, 11.0 and 23.0% of phenotypic variation. Lines within the mapping population were placed in eight categories with respect to their various QTL combinations. Lines with no QTL were the most susceptible, whereas those with the Sumai 3-derived 3BS and 6BL QTL combined with the 2DS QTL from Y1193-6 were the most resistant. Though the 3BS and 6BL QTL are well-documented, the 2DS resistance QTL, which was contributed by the susceptible parent, confers increased susceptibility when derived from Sumai 3. In this study no new FHB QTL from Sumai 3 was discovered, but results suggest that Sumai 3 contains a QTL for susceptibility on chromosome arm 2DS. Selection against this QTL may potentially increase resistance levels among Sumai 3-derived populations.

Correlation of TEM, SHV and CTX-M extended-spectrum beta lactamases among Enterobacteriaceae with their in vitro antimicrobial susceptibility

Purpose: The present study was carried out to characterize the ESBL types and evaluated their in vitro activity against a collection of Gram negative bacteria (GNB) from a multicentric Indian surveillance study. Material and Methods: During January 2005 to June 2006, six tertiary care centres in India forwarded 778 non-duplicate GNB to our reference laboratory. Three hundred GNB from this collection were selected based on clinical significance and were used in the present study. Tested isolates included Escherichia coli (167), Klebsiella spp. (122) and Enterobacter spp. (11). ESBL screening and confirmation was performed for all the isolates. Minimum inhibitory concentration of imipenem, meropenem, ertapenem, levofloxacin, amikacin, piperacillin/tazobactam and ceftriaxone was determined by the E-test method. Molecular typing of the ESBLs was performed by polymerase chain reaction among the 121 selected isolates. Results: The study showed excellent susceptibility among the strains to imipenem (100%), meropenem (100%) and ertapenem (98.7%); good susceptibility to amikacin (89.7%) and piperacillin/tazobactam (85.3%) was observed. TEM and CTX-M were predominantly found in E. coli (39.2%) while, among the Klebsiella spp., TEM, SHV and CTX-M occurred together in 42.6% of the isolates. Conclusion: More than one ESBL was produced by many strains, and this was correlated with increased resistance levels. Carbapenems continue to show good in vitro activity and ertapenem is a potential alternative to imipenem and meropenem. Continued antimicrobial resistance surveillance is warranted in light of these findings.

Face or flee? Fenitrothion resistance and behavioral response in populations of the maize weevil, Sitophilus zeamais

Insect survival in the presence of contact insecticides may be through physiological mechanisms or avoidance of contact with the compound. Curiously, although the first alternative is the object of frequent attention, the second is often neglected, but both may lead to insecticide resistance. Preliminary evidence for both physiological and behavioral resistance to pyrethroids has been obtained for a few strains of the maize weevil Sitophilus zeamais Motschulsky (Coleoptera: Curculionidae). Here we carried out a more comprehensive survey using 15 populations of S. zeamais, by examining a long-used but relatively little studied organophosphate – fenitrothion, recording not only physiological resistance, but also the behavioral responses to exposure. Physiological resistance to fenitrothion among populations of S. zeamais reached low to moderate levels (ranging from 0.9 to 14.1× at the LC 50), an increase in resistance levels compared with previous studies. Fenitrothion-induced behavioral avoidance varied among populations, particularly regarding insecticide irritability (i.e., avoidance after contact with fenitrothion), but the behavioral responses observed were mainly stimulus-independent. However, there was no correlation between physiological and behavioral resistance to fenitrothion in S. zeamais populations. Both survival strategies to fenitrothion – facing or fleeing the insecticide exposure, were observed and may co-occur in a single population, emphasizing the need of assessing both responses and their relative importance in designing management programs against stored-product insects.

Molecular Investigations of PenA-mediated β-lactam Resistance in Burkholderia pseudomallei

Burkholderia pseudomallei is the etiological agent of melioidosis. Because of the bacterium’s intrinsic resistance and propensity to establish latent infections, melioidosis therapy is complicated and prolonged. Newer generation β-lactams, specifically ceftazidime, are used for acute phase therapy, but resistance to this cephalosporin has been observed. The chromosomally encoded penA gene encodes a putative twin arginine translocase (TAT)-secreted β-lactamase, and penA mutations have been implicated in ceftazidime resistance in clinical isolates. However, the role of PenA in resistance has not yet been systematically studied in isogenetic B. pseudomallei mutant backgrounds. We investigated the effects of penA deletion, point mutations, and up-regulation, as well as tat operon deletion and PenA TAT-signal sequence mutations. These experiments were made possible by employing a B. pseudomallei strain that is excluded from Select Agent regulations. Deletion of penA significantly (>4-fold) reduced the susceptibility to six of the nine β-lactams tested and ≥16-fold for ampicillin, amoxicillin, and carbenicillin. Overexpression of penA by single-copy, chromosomal expression of the gene under control of the inducible Ptac promoter, increased resistance levels for all β-lactams tested 2- to 10-fold. Recreation of the C69Y and P167S PenA amino acid substitutions previously observed in resistant clinical isolates increased resistance to ceftazidime by ≥85- and 5- to 8-fold, respectively. Similarly, a S72F substitution resulted in a 4-fold increase in resistance to amoxicillin and clavulanic acid. Susceptibility assays with PenA TAT-signal sequence and ΔtatABC mutants, as well as Western blot analysis, confirmed that PenA is a TAT secreted enzyme and not periplasmic but associated with the spheroplastic cell fraction. Lastly, we determined that two LysR-family regulators encoded by genes adjacent to penA do not play a role in transcriptional regulation of penA expression.

Janus-activated kinases and signal transducer and activator of transcription control tumor growth response to camptothecin in human colon cancer cells

Activation of Janus kinases (JAKs) and Signal Transducers and Activators of Transcription (STATs) plays a crucial role in cell survival and proliferation. The JAK/STAT signaling pathway has received a great deal of attention as a therapeutic target for the treatment of cancer. Thus, the identification of a compound that blocks this pathway would contribute significantly to growth inhibition and apoptosis of tumor cells. The antitumor alkaloid camptothecin (CPT) may be useful in the treatment of certain cancer, but the effects of this drug on colon cancer cells remain largely undefined. The purpose of the present study was to characterize the effects of CPT on human colon cancer cells and to determine the cellular mechanisms involved in CPT-mediated cell inhibition. The cellular determinants for CPT activity were studied in six colon cancer cell lines; these cell lines exhibited natural differences in sensitivity to CPT and could be ranked according to increasing resistance levels in the order Lovo < SW48 < HCT116 < HCT8 < HT29 < WiDr. Our findings suggest that JAK2 is necessary for induction of apoptosis following CPT treatment. Inhibition of JAK2 and STAT3 Tyr705 phosphorylation decreased the expression of STAT3 downstream target genes such as Bcl-2, Bcl-xL, and Mcl-1. Finally, we show that JAK2 mRNA expression to be a better determination for CPT sensitivity than the topoisomerase-I activity or mRNA expression.