Moderate Selection Pressure Research Articles

Deciphering functional redundancy in the human microbiome

Although the taxonomic composition of the human microbiome varies tremendously across individuals, its gene composition or functional capacity is highly conserved — implying an ecological property known as functional redundancy. Such functional redundancy has been hypothesized to underlie the stability and resilience of the human microbiome, but this hypothesis has never been quantitatively tested. The origin of functional redundancy is still elusive. Here, we investigate the basis for functional redundancy in the human microbiome by analyzing its genomic content network — a bipartite graph that links microbes to the genes in their genomes. We find that this network exhibits several topological features that favor high functional redundancy. Furthermore, we develop a simple genome evolution model to generate genomic content network, finding that moderate selection pressure and high horizontal gene transfer rate are necessary to generate genomic content networks with key topological features that favor high functional redundancy. Finally, we analyze data from two published studies of fecal microbiota transplantation (FMT), finding that high functional redundancy of the recipient’s pre-FMT microbiota raises barriers to donor microbiota engraftment. This work elucidates the potential ecological and evolutionary processes that create and maintain functional redundancy in the human microbiome and contribute to its resilience.

ANALYSIS OF POLYMORPHISM IN MUCIN 4 GENE ASSOCIATED WITH ANIMAL RESISTANCE TO COLIBACTERIOSIS IN PIGS OF DOMESTIC AND FOREIGN SELECTIONS

Nowadays one of the most common and critical problem in pig breeding is colibacteriosis. This infectious disease has acute course and it is caused by enteropathogenic strains of bacteria Escherichia coli. One of the recent and effective approaches to prevent colibacteriosis is using markers of selection; it involves pig genotyping by genome loci. The chosen loci are associated with animal sensitivity to the disease and selection of animals with increased resistance by the results of genotyping. According to several researches, one of such loci is Mucin 4 (MUC4) gene. MUC4 gene is located in the13-th chromosome (SSC13q41). Mucins (MUC) are macromolecular glycoproteins synthesized by goblet enterocytes and play main role in protecting the intestinal epithelium from pathogens, including adhesive strains of Escherichia coli. As a result of spot replacement g.1849 G>С in intron 7, structure of mucin protein encoded by gene is changing which leads to changes in the sensitivity of the intestinal mucosa to pathogenic Escherichia coli. G allele and respectively GG genotype determine the resistance of animals to colibacteriosis, while СС і GС genotypes are susceptible to this disease. In Ukraine, the studies of polymorphism MUC4 g.1849 G> C were held fragmentally and only at certain populations of Ukrainian Meat and Large White. The aim of the work was to determine the genetic structure of pig breeds of domestic and foreign selection for MUC4 gene and to establish the possibility of organizing marker associated selection for genetic improvement of resistance to colibacteriosis. For research the samples of blood and hair were used from the animals of Large White of English selection, Large White of Ukrainian selection Type 1 and Type 3, Red White-Belt, Mirgorod, Poltava Meat and Landrace breeds. DNA isolation from the samples of biomaterial were carried out using ion exchange resin Chelex-100. Genotyping was performed by PCR-RFLP by Jorgensen methods (2006) with own modifications, concerning the selection of primer annealing temperature and optimum concentration of the gel to separate the restriction fragments. Genetic structure was determined using DNA analysis of MUC4 locus at seven breeds and intrabreed types. Predominance of potentially beneficial G allele frequency over undesirable C allele was established in all the populations of the studied pigs. The highest frequencies of G allele were characterized for Mirgorod (0.795), Poltava Meat (0.740) and Red White-Belt (0.820) breeds. Analysis of the distribution of genotypes showed domination of genotypes GG and GC in all analysed populations. Positive Wright fixation index and predominance of expected heterozygosity (0.484) over actual one (0.460) for Large White of Ukrainian selection Type 3 indicate the existence of moderate inbreeding and selective pressure in this herd. Other populations were characterized by a negative value of fixation index, which is indicative of an excess of heterozygotes, these breeds are in outbreeding depression. Statically significant deviation of actual frequencies of genotypes from expected ones was identified using Hardy-Weinberg criterion for pig population of Large White of English selection and Large White of Ukrainian selection Type 1. This may indicate that these populations aren’t in condition of equilibrium and about inclusion of the chosen gene to the selection process. The valuation was performed by calculating the PIC (polymorphic information content) of the marker. Based on the calculated PIC index MUC4 locus could be perspective to be used in marker associated selection with improving genetic resistance to colibacteriosis. The data of Mucin 4 gene polymorphism at Ukrainian pig populations of different origins and productive direction show the possibility of marker selection to improve the genetic resistance of animals to colibacteriosis regardless of their belonging to breed. This creates prerequisites for the establishment and implementation of the early molecular diagnosis of carriers of harmful C allele in MUC4 gene in pig breeding.

Impact of Low and High Doses of Marbofloxacin on the Selection of Resistant Enterobacteriaceae in the Commensal Gut Flora of Young Cattle: Discussion of Data from 2 Study Populations.

In the context of requested decrease of antimicrobial use in veterinary medicine, our objective was to assess the impact of two doses of marbofloxacin administered on young bulls (YBs) and veal calves (VCs) treated for bovine respiratory disease, on the total population of Enterobacteriaceae in gut flora and on the emergence of resistant Enterobacteriaceae. In two independent experiments, 48 YBs from 6 commercial farms and 33 VCs previously colostrum deprived and exposed to cefquinome were randomly assigned to one of the three groups LOW, HIGH, and Control. In LOW and HIGH groups, animals received a single injection of, respectively, 2 and 10 mg/kg marbofloxacin. Feces were sampled before treatment, and at several times after treatment. Total and resistant Enterobacteriaceae enumerating were performed by plating dilutions of fecal samples on MacConkey agar plates that were supplemented or not with quinolone. In YBs, marbofloxacin treatment was associated with a transient decrease in total Enterobacteriaceae count between day (D)1 and D3 after treatment. Total Enterobacteriaceae count returned to baseline between D5 and D7 in all groups. None of the 48 YBs harbored marbofloxacin-resistant Enterobacteriaceae before treatment. After treatment, 1 out of 20 YBs from the Control group and 1 out of 14 YBs from the HIGH group exhibited marbofloxacin-resistant Enterobacteriaceae. In VCs, the rate of fluoroquinolone-resistant Enterobacteriaceae significantly increased after low and high doses of marbofloxacin treatment. However, the effect was similar for the two doses, which was probably related to the high level of resistant Enterobacteriaceae exhibited before treatment. Our results suggest that a single treatment with 2 or 10 mg/kg marbofloxacin exerts a moderate selective pressure on commensal Enterobacteriaceae in YBs and in VCs. A fivefold decrease of marbofloxacin regimen did not affect the selection of resistances among commensal bacteria.

Evolving gene banks: improving diverse populations of crop and exotic germplasm with optimal contribution selection.

We simulated pre-breeding in evolving gene banks - populations of exotic and crop types undergoing optimal contribution selection for long-term genetic gain and management of population genetic diversity. The founder population was based on crosses between elite crop varieties and exotic lines of field pea (Pisum sativum) from the primary genepool, and was subjected to 30 cycles of recurrent selection for an economic index composed of four traits with low heritability: black spot resistance, flowering time and stem strength (measured on single plants), and grain yield (measured on whole plots). We compared a small population with low selection pressure, a large population with high selection pressure, and a large population with moderate selection pressure. Single seed descent was compared with S0-derived recurrent selection. Optimal contribution selection achieved higher index and lower population coancestry than truncation selection, which reached a plateau in index improvement after 40 years in the large population with high selection pressure. With optimal contribution selection, index doubled in 38 years in the small population with low selection pressure and 27-28 years in the large population with moderate selection pressure. Single seed descent increased the rate of improvement in index per cycle but also increased cycle time.

Synonymous codon bias and functional constraint on GC3-related DNA backbone dynamics in the prokaryotic nucleoid.

While mRNA stability has been demonstrated to control rates of translation, generating both global and local synonymous codon biases in many unicellular organisms, this explanation cannot adequately explain why codon bias strongly tracks neighboring intergene GC content; suggesting that structural dynamics of DNA might also influence codon choice. Because minor groove width is highly governed by 3-base periodicity in GC, the existence of triplet-based codons might imply a functional role for the optimization of local DNA molecular dynamics via GC content at synonymous sites (≈GC3). We confirm a strong association between GC3-related intrinsic DNA flexibility and codon bias across 24 different prokaryotic multiple whole-genome alignments. We develop a novel test of natural selection targeting synonymous sites and demonstrate that GC3-related DNA backbone dynamics have been subject to moderate selective pressure, perhaps contributing to our observation that many genes possess extreme DNA backbone dynamics for their given protein space. This dual function of codons may impose universal functional constraints affecting the evolution of synonymous and non-synonymous sites. We propose that synonymous sites may have evolved as an ‘accessory’ during an early expansion of a primordial genetic code, allowing for multiplexed protein coding and structural dynamic information within the same molecular context.

Effect of growth rate and selection pressure on rates of transfer of an antibiotic resistance plasmid between E. coli strains

Antibiotic resistance increases costs for health care and causes therapy failure. An important mechanism for spreading resistance is transfer of plasmids containing resistance genes and subsequent selection. Yet the factors that influence the rate of transfer are poorly known. Rates of plasmid transfer were measured in co-cultures in chemostats of a donor and a acceptor strain under various selective pressures. To document whether specific mutations in either plasmid or acceptor genome are associated with the plasmid transfer, whole genome sequencing was performed. The DM0133 TetR tetracycline resistance plasmid was transferred between Escherichia coli K-12 strains during co-culture at frequencies that seemed higher at increased growth rate. Modeling of the take-over of the culture by the transformed strain suggests that in reality more transfer events occurred at low growth rates. At moderate selection pressure due to an antibiotic concentration that still allowed growth, a maximum transfer frequency was determined of once per 1011 cell divisions. In the absence of tetracycline or in the presence of high concentrations the frequency of transfer was sometimes zero, but otherwise reduced by at least a factor of 5. Whole genome sequencing showed that the plasmid was transferred without mutations, but two functional mutations in the genome of the recipient strain accompanied this transfer. Exposure to concentrations of antibiotics that fall within the mutant selection window stimulated transfer of the resistance plasmid most.

Understanding Genetics of Herbicide Resistance in Weeds: Implications for Weed Management

Herbicide-resistant weeds pose a serious threat to weed management across the globe. Weeds evolve resistance to herbicides as a result of herbicide selection pressure. Under continuous herbicide selection, the resistant individuals dominate in a population. Understanding the genetics of herbicide resistance will help assess frequency and spread of herbicide resistance allele(s) in a population, which will assist in formulating prudent weed management practices to delay the evolution of resistance. Herbicide resistance, inherited by nuclear genes evolves quicker and spreads rapidly as the trait can be transmitted via pollen as opposed to maternally-inherited resistance. Dominant expression of the resistance allele also accelerates the process. Target-site resistance (TSR) is determined by a single gene, and is more likely a result of strong selection pressure. Although, non-target site resistance (NTSR) usually results under moderate selection pressure accumulating multiple alleles with minor effects imparting resistance, it may also involve a single gene when evolved under high selection pressure. Such monogenic resistances evolve and spread quickly, especially when the resistance is dominant and nuclear-inherited. Herbicide mixtures with different modes of action when applied at recommended doses can effectively delay the evolution of both TSR and NTSR.

Transgenic plants from fragmented shoot tips of apple ( Malus baccata (L.) Borkhausen) via agrobacterium-mediated transformation

Transgenic apple ( Malus baccata (L.) Borkhausen) plants were obtained via Agrobaterium-mediated transformation of fragmented shoot tips. Our results showed that without wounding treatment or with wounding treatment (II, cutting shoot tips vertically into two parts), shoots generally regenerated from meristem tissues directly and adventitious shoot regeneration was rarely observed. Otherwise, when shoot tips were cut vertically into four parts, a high percentage of callus formation (89.2%) and of adventitious shoot regeneration (60.8%) was observed. Under 20 mg l −1 kanamycin selection pressure, over 51.7% fragmented shoot tips developed callus and seven transgenic plants with GFP (Green fluorescent protein) expression were obtained from about 120 explants (efficiency of 5.8%). No transgenic plant was obtained from agrobacteria mediate transformed leaves, even though 23.2% of which formed callus after co-cultivation and selection. Molecular analysis (PCR and RT-PCR) of the transformed lines with GFP expression confirmed integration and transcription of the transgene. Under fluorescence microscopy, areas with high density of transgenic cells were observed at the cutting edges of fragmented shoot tips, which indicated that shoot regeneration from transgenic cells should be a major factor inhibiting transformation efficiency. Our experiments also showed that with moderate or low selection pressure, transgenic shoots were obtained generally accompanied by a high numbers of chimeric shoots. While by using fluorescence microscopy observation of GFP expression, the transgenic and chimeric shoots could be detected and separated precisely for further transgenic plats regeneration or multiplication. This may be very useful for apple genetic breeding, as large numbers of transgenic plants could be obtained in a short time.

Regeneration of transgenic plants from cell suspensions of transformed apple (Malus domestica Borkh. cv. Gala)

SummaryShoot regeneration in most apple cultivars is sensitive to antibiotic selection, thus Agrobacterium-mediated transformation is of low efficiency. Our experiments confirmed that, with a moderate selection pressure (10 or 20 mg l–1 kanamycin), a high percentage of callus formation (67.0% or 54.8%, respectively) was observed. Over 80.0% or 95.6% of these calli were transgenic, although the efficiency of transgenic shoot regeneration was low. Modifying the kanamycin selection pressure, or shortening the time of kanamycin selection to 2 weeks, did not increase the efficiency of transformation. Multiplying suspension cultures could be produced from transgenic calli using single cells or small cell aggregates to initiate such cultures. When these transgenic cell aggregates were placed on regeneration medium without kanamycin selection, 17.3% of them regenerated shoots after 8 weeks, and at least 86 transgenic plants were obtained from one suspension culture line. This level of success makes it possible to obtain large numbers of transgenic apple plants from a single experiment for genetic breeding, and to study the mechanisms of integration of target genes.

Elucidating acetate tolerance in E. coli using a genome-wide approach

Engineering organisms for improved performance using lignocellulose feedstocks is an important step towards a sustainable fuel and chemical industry. Cellulosic feedstocks contain carbon and energy in the form of cellulosic and hemicellulosic sugars that are not metabolized by most industrial microorganisms. Pretreatment processes that hydrolyze these polysaccharides often also result in the accumulation of growth inhibitory compounds, such as acetate and furfural among others. Here, we have applied a recently reported strategy for engineering tolerance towards the goal of increasing Escherichia coli growth in the presence of elevated acetate concentrations (Lynch et al., 2007). We performed growth selections upon an E. coli genome library developed using a moderate selection pressure to identify genomic regions implicated in acetate toxicity and tolerance. These studies identified a range of high-fitness genes that are normally involved in membrane and extracellular processes, are key regulated steps in pathways, and are involved in pathways that yield specific amino acids and nucleotides. Supplementation of the products and metabolically related metabolites of these pathways significantly increased growth rate (a 130% increase in specific growth) at inhibitory acetate concentrations. Our results suggest that acetate tolerance will not involve engineering of a single pathway; rather we observe a range of potential mechanisms for overcoming acetate based inhibition.

Selecting survivors in genetic algorithm using tabu search strategies

Genetic algorithm (GA) is well-known for its effectiveness in global search and optimization. To balance selection pressure and population diversity is an important issue of designing GA. This paper proposes a novel hybridization of GA and tabu search (TS) to address this issue. The proposed method embeds the key elements of TS—tabu restriction and aspiration criterion—into the survival selection operator of GA. More specifically, the tabu restriction is used to prevent inbreeding for diversity maintenance, and the aspiration criterion is activated to provide moderate selection pressure under the tabu restriction. The interaction of tabu restriction and aspiration criterion enables survivor selection to balance selection pressure and population diversity. The experimental results on numerical and combinatorial optimization problems show that this hybridization can significantly improve GAs in terms of solution quality as well as convergence speed. An empirical analysis further identifies the influences of the TS strategies on the performance of this hybrid GA.

Biodegradation of acetonitrile by adapted biofilm in a membrane-aerated biofilm reactor

A membrane-aerated biofilm reactor (MABR) was developed to degrade acetonitrile (ACN) in aqueous solutions. The reactor was seeded with an adapted activated sludge consortium as the inoculum and operated under step increases in ACN loading rate through increasing ACN concentrations in the influent. Initially, the MABR started at a moderate selection pressure, with a hydraulic retention time of 16 h, a recirculation rate of 8 cm/s and a starting ACN concentration of 250 mg/l to boost the growth of the biofilm mass on the membrane and to avoid its loss by hydraulic washout. The step increase in the influent ACN concentration was implemented once ACN concentration in the effluent showed almost complete removal in each stage. The specific ACN degradation rate achieved the highest at the loading rate of 101.1 mg ACN/g-VSS h (VSS, volatile suspended solids) and then declined with the further increases in the influent ACN concentration, attributed to the substrate inhibition effect. The adapted membrane-aerated biofilm was capable of completely removing ACN at the removal capacity of up to 21.1 g ACN/m(2) day, and generated negligible amount of suspended sludge in the effluent. Batch incubation experiments also demonstrated that the ACN-degrading biofilm can degrade other organonitriles, such as acrylonitrile and benzonitrile as well. Denaturing gradient gel electrophoresis studies showed that the ACN-degrading biofilms contained a stable microbial population with a low diversity of sequence of community 16S rRNA gene fragments. Specific oxygen utilization rates were found to increase with the increases in the biofilm thickness, suggesting that the biofilm formation process can enhance the metabolic degradation efficiency towards ACN in the MABR. The study contributes to a better understanding in microbial adaptation in a MABR for biodegradation of ACN. It also highlights the potential benefits in using MABRs for biodegradation of organonitrile contaminants in industrial wastewater.

Diversity and evolution of the small multidrug resistance protein family

BackgroundMembers of the small multidrug resistance (SMR) protein family are integral membrane proteins characterized by four α-helical transmembrane strands that confer resistance to a broad range of antiseptics and lipophilic quaternary ammonium compounds (QAC) in bacteria. Due to their short length and broad substrate profile, SMR proteins are suggested to be the progenitors for larger α-helical transporters such as the major facilitator superfamily (MFS) and drug/metabolite transporter (DMT) superfamily. To explore their evolutionary association with larger multidrug transporters, an extensive bioinformatics analysis of SMR sequences (> 300 Bacteria taxa) was performed to expand upon previous evolutionary studies of the SMR protein family and its origins.ResultsA thorough annotation of unidentified/putative SMR sequences was performed placing sequences into each of the three SMR protein subclass designations, namely small multidrug proteins (SMP), suppressor of groEL mutations (SUG), and paired small multidrug resistance (PSMR) using protein alignments and phylogenetic analysis. Examination of SMR subclass distribution within Bacteria and Archaea taxa identified specific Bacterial classes that uniquely encode for particular SMR subclass members. The extent of selective pressure acting upon each SMR subclass was determined by calculating the rate of synonymous to non-synonymous nucleotide substitutions using Syn-SCAN analysis. SUG and SMP subclasses are maintained under moderate selection pressure in comparison to integron and plasmid encoded SMR homologues. Conversely, PSMR sequences are maintained under lower levels of selection pressure, where one of the two PSMR pairs diverges in sequence more rapidly than the other. SMR genomic loci surveys identified potential SMR efflux substrates based on its gene association to putative operons that encode for genes regulating amino acid biogenesis and QAC-like metabolites. SMR subclass protein transmembrane domain alignments to Bacterial/Archaeal transporters (BAT), DMT, and MFS sequences supports SMR participation in multidrug transport evolution by identifying common TM domains.ConclusionBased on this study, PSMR sequences originated recently within both SUG and SMP clades through gene duplication events and it appears that SMR members may be evolving towards specific metabolite transport.

Genetic variation and differentiation of floral morphology in wild Primula sieboldii evaluated by image analysis data and SSR markers

Primula sieboldii (E. Morren), the ancestor of the Japanese garden Primula, is in danger of extinction in the wild. Genetic diversity is a key component for conservation efforts associated with population management. Genetic diversity in visible traits and several molecular makers were evaluated, respectively. Since it is difficult to determine the degree to which genetic and environmental differences contribute to observed phenotypic variation in natural habitats. A common-garden approach is used. The contribution of genetic and environmental differences to variation in petal shape and area of Primula sieboldii were evaluated. Samples from 108 genets gathered from five natural populations in three regions of Japan were analyzed in a common-garden experiment and also analyzed using eight microsatellite markers. From the results of quantitative evaluation based on image analysis, broad genetic variation in petal traits within populations and low level of population differentiation was found. For all petal shapes, Qst was smaller than Fst, suggesting that wild populations might be under moderate selective pressure for a specific phenotype. For petal area, Qst was nearly equal to Fst, suggesting that population differentiation has been caused mainly by genetic drift.

Additive genetic variance for stem strength in field pea (Pisum sativum)

Weak stem strength in field pea (Pisum sativum) is a major restriction to yield, seed quality and ease of harvest. Three aspects of stem strength: load at breaking point, flexion and compressed stem thickness, showed substantial genetic variation among a diverse range of six parents including modern cultivars, landrace accessions, and interspecific progeny. Diallel analysis of parents and F1 progeny was conducted using a simple additive-dominance model, which was adequate for load and compressed stem thickness. There were significant additive genetic effects for load and compressed stem thickness with no evidence of dominance or maternal effects, and also significant additive genetic effects for flexion which was subject to more complex genetic control. Valuable alleles for these stem strength traits were present in commercial cultivars and landrace types of field pea. Efficient and practical breeding for improved stem strength will involve several recurrent selection cycles with moderate selection pressure for compressed stem thickness in early generations, followed by verification of improvements in lodging resistance in subsequent field trials. Compressed stem thickness is relatively easy to measure on individual plants in the field and is closely associated with load.

Start-Up, Microbial Community Analysis and Formation of Aerobic Granules in a tert-Butyl Alcohol Degrading Sequencing Batch Reactor

We demonstrate that compact well-settling aerobic granules can be developed in a sequencing batch reactor (SBR) for the biological removal of tert-butyl alcohol (TBA) using a strategy involving step increases in TBA loading rate achieved through increasing TBA concentrations in the influent. A moderate selection pressure that included a cycle time of 24 h and a start-of-cycle TBA concentration of 100 mg/L was initially introduced to encourage the growth and retention of biomass and avoid biomass loss from hydraulic washout. Start-of-cycle TBA concentrations were increased to 150, 300, 450, and 600 mg/L on days 90, 100, 121, and 199, respectively. These increases were only introduced after complete TBA removal was accompanied by visible improvements in biomass concentration and biomass settling ability. This acclimation strategy produced incrementally higher biomass concentrations and better settling biomass with higher specific TBA biodegradation rates. Effluent TBA concentrations were consistently below the detection limit of 25 microg/L. Aerobic granules were first observed about 180 days after reactor start-up. The granules had a clearly defined shape and appearance, settled significantly faster than the suspended sludge in the reactor, and eventually became the dominant form of biomass in the reactor. The adapted granules were capable of complete TBA removal and contained a stable microbial population with a low diversity of sequences of community 16S rRNA gene fragments. This study indicates that it is possible to use aerobic granules for TBA remediation and will contribute to a better understanding of how microbial acclimation can be exploited in the SBR to biologically remove recalcitrant xenobiotics.

Formation of granules and Methanosaeta fibres in an anaerobic migrating blanket reactor (AMBR).

It has generally been accepted that the formation of granules in anaerobic wastewater treatment systems requires a hydraulic upflow pattern. To evaluate this hypothesis, we operated an anaerobic migrating blanket reactor (AMBR) without a hydraulic upflow pattern, using a synthetic wastewater containing acetate, propionate, butyrate and sucrose. We provided conditions amenable to the formation of granules by operating the system with a moderate hydraulic selection pressure, which in this system was not the result of a hydraulic upflow pattern, but was provided by migration of biomass and intermittent mechanical mixing. Granules were first noticed after 2 months of operation, and it took another 2 months for a mature granular blanket to develop. Besides granules, approximately 1-cm-long Methanosaeta fibres developed and, after 6 months of operation, 30% of biomass consisted of these fibres. Quantitative membrane hybridization showed that almost all the total 16S rRNA extracted from fibres consisted of 16S rRNA from Methanosaeta concilii. This finding indicates that it was possible to develop pockets consisting almost entirely of an organism with a very limited substrate utilization spectrum (only acetate) in a system that was fed a synthetic wastewater containing acetate, propionate, butyrate and sucrose and that is known for its ability to develop biomass with a complex microbial community structure.

On the harmonious mating strategy through tabu search

Genetic algorithms (GAs) are well-known heuristic algorithms and have been applied to solve a variety of complicated problems. When adopting GA approaches, two important issues––selection pressure and population diversity––must be considered. This work presents a novel mating strategy, called tabu genetic algorithm (TGA), which harmonizes these two issues by integrating tabu search (TS) into GA’s selection. TGA incorporates the tabu list to prevent inbreeding so that population diversity can be maintained, and further utilizes the aspiration criterion to supply moderate selection pressure. An accompanied self-adaptive mutation method is also proposed to overcome the difficulty of determining mutation rate, which is sensitive to computing performance. The classic traveling salesman problem is used as a benchmark to validate the effectiveness of the proposed algorithm. Experimental results indicate that TGA can achieve harmony between population diversity and selection pressure. Comparisons with GA, TS, and hybrids of GA and TS further confirm the superiority of TGA in terms of both solution quality and convergence speed.

Susceptibility of Listeria monocytogenes and Listeria innocua strains isolated from short-ripened cheeses to some antibiotics and heavy metal salts

Twenty-eight isolates of Listeria monocytogenes and 18 of L. innocua obtained from short- ripened cheeses manufactured in Asturias (northern Spain) were screened for antibiotic and heavy metal salt susceptibility patterns. No remarkable differences were detected between L. monocytogenes and L. innocua or between clusters previously defined in each species by pulsed-field gel electrophoresis analysis of the chromosomal DNA. β -lactams, tetracycline, erythromycin and rifampicin were the most effective antibiotics with MIC (minimum inhibitory concentration) variations within the species not exceeding one order of dilution. Chloramphenicol, streptomycin, kanamycin, gentamicin and neomycin displayed poor activity and their MICs showed a wider range of values than the other antibiotics. Significant rates of low-level resistance to streptomycin, kanamycin and gentamicin were also recorded. Fosfomycin was not effective againstListeria as the isolates were resistant to high levels of this antibiotic. All strains displayed high susceptibility to mercuric chloride and were resistant to high lead nitrate concentrations. Susceptibility to ferric citrate, cupric, zinc and cadmium sulfate was intermediate between that of mercuric chloride and lead nitrate. All isolates had similar susceptibility patterns against mercury, iron, zinc and lead salts. However, one L. monocytogenes strain showed lower tolerance to cupric sulfate than the remaining population and the MICs for cadmium sulfate varied in a wider range of concentrations than for the other salts. Clearly differentiated resistant strains with respect to a more sensitive population were not detected for any antimicrobial agent. These data suggest a moderate selective pressure exerted by aminoglycosides, cadmium and cupric sulfate but not by other antimicrobials in dairy farm environments of Listeria isolates tested.

Efficacy of Bacillus thuringiensis Serovar Israelensis against Bacillus sphaericus Resistant and Susceptible Larvae of Culex quinquefasciatus Say

A potential biological larvicides like Bacillus sphaericus (Bs) has been advocated in mosquito control operations. However, due to rapid development of resistance of mosquitoes to Bs toxin alternate mosquito control measures are needed. A strain of Culex quinquejasciatus Say was found highly resistant (resistance ratio (RR) = 6223 folds) in the field (Gandhinagar, Kochi, Kerala) during the year 1995 and was reared in the laboratory by subjecting to moderate selection pressure using Bs 2362 strain to maintain resistance. A susceptible mosquito colony of C. quinquejasciatus was also reared simultaneously. Bioassays were conducted with lyophilized bacterial culture of spore/crystal toxin of Bs 2362 with the inbred generations obtained from these colonies. We found that a high level of resistance to Bs (2362) toxin (RR at LC 50 = 249.2 and at LC 90 Bacillus thuringiensis serovar israelensis (BIi) (VeetoBac*). This observation suggest that Bli based biopesticide can be used for the management of Bs resistant mosquito control operations.