Average Generation Time Research Articles

Growth of Aeromonas hydrophila in the Whey Cheeses Myzithra, Anthotyros, and Manouri during Storage at 4 and 12°C

The fresh whey cheeses Myzithra, Anthotyros, and Manouri were inoculated with Aeromonas hydrophila strain NTCC 8049 (type strain) or with an A. hydrophila strain isolated from food (food isolate) at levels of 3.0 to 5.0 × 102 CFU/g of cheese and stored at 4 or 12°C. Duplicate samples of cheeses were tested for levels of A. hydrophila and pH after up to 29 days of storage. At 4°C, A. hydrophila grew in Myzithra and Anthotyros with a generation time of ca. 19 h, but no growth was observed in Manouri. In Myzithra, average maximum populations of 8.87 log CFU/g (type strain) and 8.79 log CFU/g (food isolate) were recorded after 20 and 22 days of storage at 4°C, respectively. The average maximum populations observed in Anthotyros stored at 4°C were 6.72 log CFU/g (food isolate) and 6.13 log CFU/g (type strain) and were observed after 15 and 16 days of storage, respectively. A. hydrophila grew rapidly and reached high numbers in cheeses stored at 12°C. The average generation times were 3.7 and 3.9 h (Myzithra), 4.1 and 6.1 h (Anthotyros), and 8.0 and 9.2 h (Manouri) for the type strain and the food isolate, respectively. Among the different whey cheese trials, the highest A. hydrophila population recorded (10.13 log CFU/g) was in Myzithra that had been inoculated with the food isolate after 8 days of storage at 12°C. To prevent A. hydrophila growth in whey cheeses, efforts must be focused on preventing postprocessing contamination and temperature abuse during transportation and storage.

SLIM: an alternative Web interface for MEDLINE/PubMed searches – a preliminary study

BackgroundWith the rapid growth of medical information and the pervasiveness of the Internet, online search and retrieval systems have become indispensable tools in medicine. The progress of Web technologies can provide expert searching capabilities to non-expert information seekers. The objective of the project is to create an alternative search interface for MEDLINE/PubMed searches using JavaScript slider bars. SLIM, or Slider Interface for MEDLINE/PubMed searches, was developed with PHP and JavaScript. Interactive slider bars in the search form controlled search parameters such as limits, filters and MeSH terminologies. Connections to PubMed were done using the Entrez Programming Utilities (E-Utilities). Custom scripts were created to mimic the automatic term mapping process of Entrez. Page generation times for both local and remote connections were recorded.ResultsAlpha testing by developers showed SLIM to be functionally stable. Page generation times to simulate loading times were recorded the first week of alpha and beta testing. Average page generation times for the index page, previews and searches were 2.94 milliseconds, 0.63 seconds and 3.84 seconds, respectively. Eighteen physicians from the US, Australia and the Philippines participated in the beta testing and provided feedback through an online survey. Most users found the search interface user-friendly and easy to use. Information on MeSH terms and the ability to instantly hide and display abstracts were identified as distinctive features.ConclusionSLIM can be an interactive time-saving tool for online medical literature research that improves user control and capability to instantly refine and refocus search strategies. With continued development and by integrating search limits, methodology filters, MeSH terms and levels of evidence, SLIM may be useful in the practice of evidence-based medicine.

Cell cycle and centromere FISH studies in premature centromere division

BackgroundMitotic configurations consistent in split centromeres and splayed chromatids in all or most of the chromosomes or premature centromere division (PCD) have been described in three categories. (1) Low frequency of PCD observed in colchicines-treated lymphocyte cultures from normal individuals. (2) High frequency of PCD with mosaic variegated aneuploidy. (3) High frequency of PCD as a sole chromosome abnormality observed in individuals with no recognizable clinical pattern. We report four members of a family with the third category of PCD.MethodsCell cycle duration assessed by average generation time using differential sister chromatid stain analysis and FISH studies of DNA centromere sequences in PCD individuals, are included and compared with previously reported PCD individuals from 9 families.ResultsWe observed PCD in colchicine-treated cultures from the propositus, his father, and two paternal aunts but not in his mother and four other paternal and maternal family members, as well as in untreated cultures from the propositus and his father. We observed cytological evidence of active centromeres by Cd stain. Significative cell cycle time reduction in anaphases of PCD individuals (average generation time of 21.8 h;SD 0.4) with respect to individuals without PCD (average generation time of 31.8 h;SD 3.9) was observed (P < 0.005, Student t-test for independent samples). Increased cell proliferation kinetics was observed in anaphasic cells of individuals with PCD, by differential sister chromatid stain analysis. FISH studies revealed the presence of alpha satellite DNA from chromosomes 1, 13, 21/18, X, all centromeres, and CENP-B box sequences in metaphasic and anaphasic cells from PCD individuals.ConclusionThis report examines evidences of a functional relationship between PCD and cell cycle impairment. It seems that essential centromere integrity is present in these cases.

Biology and Life History of &lt;I&gt;Ascia monuste monuste&lt;/I&gt; (Lepidoptera: Pieridae), a Potential Pest of Cruciferous Vegetables

Abstract Ascia monuste (L.) (Lepidoptera: Pieridae) is a subtropical and tropical species with several subspecies and forms. In recent years, it has become a sporadic insect pest of crucifer vegetables in southern Texas. In some isolated fields, the gregarious larvae have completely consumed all aboveground parts of large cabbage or collard plants (leaves, petioles, and stems). To understand more about this subspecies, I studied its biology on cabbage, including the development of immature stages, the longevity of adults, and other parameters in the laboratory and the greenhouses. At 25 ± 2°C, developmental time averaged 4.7, 2.8, 2.3, 3.1, 3.2, 6.0, and 9.d d for eggs, first, second, third, fourth, fifth instars and pupae, respectively, yielding an average of 17.4 d for all larval stages and an average generation time of 31.7 d for both sexes. Adult longevity averaged 19.8 d for females and 16.3 d for males under greenhouse conditions. The sex ratio was 1:0.76 (females:males) or 56.8% females. Each egg c...

Interactive optical trapping shows that confinement is a determinant of growth in a mixed yeast culture

Applying a newly developed user-interactive optical trapping system, we controllably surrounded individual cells of one yeast species, Hanseniaspora uvarum, with viable cells of another yeast species, Saccharomyces cerevisiae, thus creating a confinement of the former. Growth of surrounded and non-surrounded H. uvarum cells was followed under a microscope by determining their generation time. The average generation time of surrounded H. uvarum cells was 15% higher than that of non-surrounded cells, thereby showing that the confinement imposed by viable S. cerevisiae cells on H. uvarum inhibits growth of the latter. This study is the first to demonstrate that confinement is a determinant of growth in a microbial ecosystem.

Sanguinarine: an evaluation of in vivo cytogenetic activity

Sanguinarine (SG), a benzophenanthridine alkaloid, has been shown to possess anti-microbial, anti-inflammatory and antioxidant properties. In the UK and USA its salts has been in use in mouthwashes and toothpastes to inhibit dental plaque and improve gingival health. In India and Nepal consumption of mustard oil contaminated with argemone seeds containing sanguinarine, was associated with “dropsy” syndrome. In the present study, SG was evaluated in vivo in mouse bone marrow cells for its ability to induce clastogenicity and DNA damage in terms of increased sister chromatid exchange (SCE) frequencies. Doses of 5, 10, and 15 mg/kg body weight of SG given intra peritoneally induced a positive dose-dependent significant clastogenicity and SCE frequency increases (trend test α ≤ 0.05). The minimum effective concentration to induce clastogenic and DNA damage was 10 mg of SG/kg body weight. In addition to examining SCEs, the BrdUrd-differential technique was utilized to assess the effect of SG on cell replication. The analysis revealed that SG treatment did not significantly affect the distribution of cells among the different phases of the cell cycle. The proliferation rate index and average generation time data were statistically non-significant. This indicated that the alkaloid was not cytotoxic to the bone marrow cells at the doses tested. Based on the results of the present findings, the use of this alkaloid should be restricted.

Estimates of Intracellular Delay and Average Drug Efficacy from Viral Load Data of HIV-Infected Individuals under Antiretroviral Therapy

We analyse viral load data of five patients under ritonavir monotherapy using a model of HIV dynamics under antiretroviral therapy that includes both drug pharmacokinetics and the intracellular delay from the time of cell infection to viral production. Using this approach we separate pharamacokinetic from intracellular delays, and obtain new estimates of intracellular delay and the antiviral efficacy of ritonavir. We find the average intracellular delay to be 1 day, in agreement with experiments. The average viral generation time is now estimated at 2 days, resulting in approximately 180 replication cycles per year. The model also reveals that ritonavir monotherapy is approximately 65% as efficacious as a recently used potent four-drug therapy, suggesting that selection for drug resistance may be facilitated by the relatively low efficacy of individual drugs, contributing in part to the inherent limitations of current therapies in combating HIV-1 infection.

Use of a quantitative gene expression assay based on micro-array techniques and a mathematical model for the investigation of chlamydial generation time

Chlamydia is an important pathogen which possesses a unique developmental cycle. We used real-time PCR technology to measure gene transcript levels in Chlamydia trachomatis strain L2. By measuring 16S rRNA transcript levels, and developing a mathematical model of the chlamydial developmental cycle fitting the data, we predict an average generation time of approximately 2.6 h. Additionally, potentially this modelling also provides the foundation for the application of emerging micro-array technology in which identification of the gene signals that trigger a chlamydial body to start replicating or transform to its infectious form can be made possible.

Effect of temperature on development and reproduction in Gastrophysa viridula (Coleoptera: Chrysomelidae)

The duration of development, reproduction and longevity of Gastrophysa viridula (DeGeer) was measured at constant temperatures and a long day photoperiod. At 18, 21.5, 25, and 28°C the average duration of development of the egg, larval and pupal stages and total development time (28.2, 21.6, 16.1, 15.0 days) decreased with temperature but the proportion of time spent in the egg, larval and pupal stages did not significantly change with temperature. Total development required 304.6 day degrees above the lower development threshold of 7.1°C. Pre-adult mortality and the rate of oviposition increased, and the duration of oviposition decreased with increasing temperature. Net reproduction rate Ro decreased (from 157 female eggs at 18°C to 75 female eggs at 28°C) and mean generation time T also decreased (from 45.5 days at 18°C to 24.1 days at 28°C) with increasing temperature. The intrinsic rate of population increase rm increased with temperature (from 0.111 at 18°C to 0.179 at 28°C). On a physiological time scale the average generation time T was 496 day degrees.

Effects of chlorophyllin on acetaldehyde: lack of modulation of the rate of sister-chromatid exchanges in mouse bone marrow, and of complex formation in aqueous solution

Acetaldehyde (Ace) is a reactive compound widely found in natural and industrialized products. On the other hand, chlorophyllin (Chl) is a chloropyll derivative which has shown DNA modulatory effects in several models. The first aim of the present study was to determine the capacity of Ace to increase the rate of sister-chromatid exchanges (SCEs) in mouse bone marrow cells in vivo, as well as to determine its capacity to modify the mitotic index (MI) and the average generation time (AGT). For this experiment we tested four dosages of Ace by the ip route (0.4, 4.0, 40.0 and 400 mg/kg), and found a genotoxic effect with the two highest dosages (more than double the basal level was observed with 400 mg/kg). We also found that none of the doses tested modified the MI or the AGT. A second objective was to explore the potential of Chl to modulate the genotoxicity of Ace in the same model. We evaluated whether an oral administration of Chl (2.0, 6.0 and 10.0 mg/kg), given 1 h before an ip administration of Ace (100 mg/kg), could modulate the SCEs produced by the mutagen. The result showed a similar SCE rate in both, the Ace-treated mice and those administered with the two chemicals, indicating that Chl was not a modulatory chemical on the genotoxicity of Ace. No modifications were observed concerning the MI or the AGT either. A third objective was to determine whether the two compounds (Ace and Chl) may form a molecular complex in aqueous solution. In agreement with the lack of modulatory effect by Chl, a reversed HPLC and a spectrophotometric analysis showed that the two compounds were unable to form a complex. This report confirms the importance of the specificity concerning the interaction mutagen/antimutagen.

Griseofulvin induces mitotic delay and aneuploidy in bone marrow cells of orally treated mice.

Griseofulvin (GF) is a fungicide drug well characterized for its aneugenic activity in vitro. In vivo strong evidence of aneuploidy and polyploidy induction has been obtained in germ cells, especially in oocytes. More controversial are the data on the aneugenicity of griseofulvin in somatic cells. In this paper we provide evidence that GF induces non-disjunction and cell cycle delay in bone marrow cells of orally treated mice. Adult female mice were administered olive oil suspensions of 200, 666 or 2000 mg/kg GF by gavage and killed 18 or 24 h later. To minimize animal-to-animal variation in the absorption and distribution of GF, mice were fasted from the time of GF administration to the time of killing. Two hours before treatment the animals were s.c. implanted with a bromodeoxyuridine tablet to obtain differential chromatid staining and to determine the number of divisions after GF treatment for each metaphase. Mitostatic effects of GF were assessed by the relative proportions of first, second and third generation metaphases and the average generation time (AGT) method. A statistically significant increase with respect to the control AGT value was observed after treatment with 666 and 2000 mg/kg, suggesting that GF, as already shown in meiosis, interfered with cell cycle progression. Hyperploidy was scored in second generation metaphases. Eighteen hours after treatment, the frequencies of hyperploid cells were significantly (P < 0.05) higher in all GF-treated groups than in their matched control group. The effect was not dose-dependent. No further increase in aneuploidy was observed at 24 h, suggesting that cells overcoming mitotic arrest did not have a higher rate of non-disjunction. No induction of polyploidy was demonstrated. We conclude that GF induces mitotic delay and aneuploidy in mouse bone marrow and suggest that the protocol used to formulate the gavage suspensions and the after-treatment fasting of the animals enhanced the bioavailability of GF to bone marrow cells.

Competition between Moina macrocopa and Ceriodaphnia dubia: a Life Table Demography StudyPart of the data presented at the International Plankton Meet, Chapala (Mexico)

We analyzed the effect of competition on survival- and reproduction-related parameters under two (0.5.10 6 and 1.5.10 6 cells ml -1 of Chlorella) food densities between Moina macrocopa and Ceriodaphnia dubia using the life table demography approach. In general, regardless of the presence of Moina, the average lifespan and generation time were higher for C. dubia. However, rate of population increase (r) and values of reproductive effort were higher for M. macrocopa. The net reproductive values of C. dubia grown alone varied from 7.6 to 36.6 offspring female 1 lifetime -1 , depending on Chlorella density. Corresponding values for Moina varied little (16.4 and 17.4 offspring female -1 lifetime -1 ) Values of gross reproductive rate, net reproductive rate and the generation time of both cladoceran species were higher in mixed cultures than when grown alone. The negative effect of competition was evident for both the cladoceran species, not in their reproductive rates, but in their generation times, and thus population growth rates. The role of competition and facilitation occurring between the two species was discussed in relation to different algal food levels.

Genotoxic effects of malathion: an organophosphorus insecticide, using three mammalian bioassays in vivo

The genotoxic effects of malathion was evaluated using chromosome aberration, sister chromatid exchange (SCE) and sperm abnormality assays in mice. All the three acute doses (2.5, 5 and 10 mg/kg) of malathion tested in the present study, induced significant dose-dependent increase in the frequency of chromosome aberrations and sperm abnormalities, but did not affect the total sperm count. The highest acute dose induced a >12-fold increase in the frequency of chromosome aberrations, two-fold increase in the frequency of SCEs and four-fold increase in the frequency of sperms with abnormal head morphology following intraperitoneal (i.p.) exposure. Further, a significant increase in the frequency of SCEs was observed, but the increase was not dose-dependent. At higher doses, malathion induced a moderate delay in cell cycle as evident from the increase in average generation time (AGT). The present findings suggest that technical grade malathion is a potent genotoxic agent and may be regarded as a potential germ cell mutagen also.

Differential sensitivity of Chinese hamster V79 and Chinese hamster ovary (CHO) cells in the in vitro micronucleus screening assay

Both the V79 and CHO cell lines are routinely used in the in vitro MN screening assay for the detection of possible genotoxicants. The CHO cell line is the predominant cell line currently used in the genetic toxicology testing industry. However, some laboratories routinely utilize the V79 cell line since the in vitro MN screening assay was initially developed using V79 cells. Our laboratory has historically used the CHO cell line. Therefore, our laboratory was interested in comparing the two cell lines with regard to possible similarities or differences in MN induction sensitivity after exposure to cyclophosphamide (CPA) and mitomycin C (MMC), the two standard positive control chemicals routinely used in this assay. Three exposure conditions in the presence of CPA and MMC were examined in both cell lines. Replicate cultures of CHO cells in McCoy’s 5A and V79 cells in both McCoy’s 5A and E-MEM were established and treated with 5 μg CPA/ml (4 h exposure with S9), 0.5 μg MMC (4 h exposure without S9) and 0.5 μg MMC (24 h exposure without S9). A total of 400 cytochalasin B-blocked binucleated cells and 200 consecutive cells were analyzed from each culture for MN and cell cycle kinetics, respectively. Analysis of the data demonstrated that CHO cells were up to approximately five-fold more sensitive to the induction of CPA- and MMC-induced MN than V79 cells. Both cell lines exhibited similar average generation times among identical exposure groups. Therefore, the difference in MN sensitivity cannot be attributed to possible differences in cell cycle kinetics and is possibly related to inherent cellular differences in the processing of and/or repair of CPA- and MMC-induced damage by V79 and CHO cells.

Lack of genetic differentiation among widely spaced subpopulations of a butterfly with home range behaviour.

We examine seven geographically separate subpopulations of Heliconius charithonia, a butterfly with well-documented home range behaviour, in Miami-Dade County, Florida, for genetic differentiation using cellulose acetate electrophoresis. These subpopulations exhibit little genetic variation (percent polymorphic loci = 27, average heterozygosity = 0.103) especially in comparison to populations of the same and related species from mainland South America. Allele frequencies do not differ among the subpopulations in south Florida and estimates of Wright's fixation index (FST) support that there is no detectable genetic differentiation among them. This result supports an earlier finding that the dispersal ability of Heliconius butterflies may be underestimated. However, it is unlikely that increased dispersal ability alone could account for the lack of genetic differentiation observed among subpopulations separated by almost 80 km. Given the likely effective population size of these subpopulations (Ne = 205) and the average generation time of this species in the subtropics (in the range of 30-90 days), this lack of genetic differentiation is best explained by current or very recent gene flow following a stepping-stone model. Furthermore, this result provides evidence that the current extensive degree of habitat fragmentation surrounding the city of Miami does not limit gene flow among urban subpopulations of Heliconius charithonia.

Circadian clock-protein expression in cyanobacteria: rhythms and phase setting.

The cyanobacterial gene cluster kaiABC encodes three essential circadian clock proteins: KaiA, KaiB and KaiC. The KaiB and KaiC protein levels are robustly rhythmical, whereas the KaiA protein abundance undergoes little if any circadian oscillation in constant light. The level of the KaiC protein is crucial for correct functioning of the clock because induction of the protein at phases when the protein level is normally low elicits phase resetting. Titration of the effects of the inducer upon phase resetting versus KaiC level shows a direct correlation between induction of the KaiC protein within the physiological range and significant phase shifting. The protein synthesis inhibitor chloramphenicol prevents the induction of KaiC and blocks phase shifting. When the metabolism is repressed by either translational inhibition or constant darkness, the rhythm of KaiC abundance persists; therefore, clock protein expression has a preferred status under a variety of conditions. These data indicate that rhythmic expression of KaiC appears to be a crucial component of clock precession in cyanobacteria.

Influence of disinfection processes on the microbial quality of potable groundwater in a laboratory-scale system model

Groundwater was used to evaluate the influence of the disinfection processes on the microbial quality of potable water distribution systems using laboratory-scale units. Coliform bacteria, heterotrophic plate count and total bacteria were used for the evaluation of the bactericidal effectiveness of each disinfectant. The microbial disinfection efficacy of chlorine, chloramine, ozone and UV irradiation was found to be equally effective in the elimination of coliform bacteria during the first hours (20 min-2 h) after disinfection. Complete elimination of coliforms in hydrogen peroxide treated water occurred after 48 h. More than 4 log cfu . ml −1 (average killing rate) heterotrophic bacteria were killed by all the disinfectants with the exception of hydrogen peroxide (average killing rates: 3-2 log cfu . ml −1 ). However, ozone was highly effective within the first 2 h as shown by the average killing rate of 4 log cfu . ml −1 heterotrophic bacteria in both source waters. The phenomenon of bacterial regrowth was linked to the absence of concentrations of disinfectant residuals. Bacterial regrowth, however, could be detected earlier with chlorine (after 20 min-average regrowth rate 0.064 h −1 , average generation time 10.95 h), ozone (after 2 h-average regrowth rate 0.202 h −1 , average-generation time 5.04 h) and UV treated water (after 2 h-average regrowth rate 0.263 h −1 , average generation time 2.70 h) than chloramine (between 24 h and 48 h-average regrowth rate 0.057 h −1 , average generation time 13.87 h) and hydrogen peroxide treated water (after 48 h-average regrowth rate 0.063 h −1 , average generation time 12.74 h). The greater persistence of monochloramine (7 days) and hydrogen peroxide (14 days) residuals were found to inhibit bacterial regrowth in potable water.

Phospholipase D in Tetrahymena: activity and significance

We detected phospholipase D in three species of ciliates: Tetrahymena: T. thermophila, T. pyriformis and T. setosa in nutrient medium supplemented with ethanol in in vivo systems, by the appearance of phosphatidylethanol. The calcium ionophore A23187 increased the synthesis of phosphatidylethanol, as compared with untreated controls. We suggest that Tetrahymena possess a calcium sensitive phospholipase D. Propranolol caused the cells in dense cultures to increase their average generation times or die, dependent on the drug concentration. This inhibition could be overcome by the addition of phospholipids or ethanol. Pure phosphatidylethanol had no effect on growth rates or generation times in cultures at high cell density, but postponed cell death in cultures at low cell density by a factor of 10. We suggest that an important role of phospholipase D in Tetrahymena is to supply the cell with diacylglycerol without which it can not enter the mode of proliferation from the lag phase of the culture.

Effects of tetramethylthiuram disulphide (thiram) on adenine nucleotide (ATP, ADP, AMP) levels in the ciliate Tetrahymena pyriformis

Axenic cultures of Tetrahymena pyriformis GL populations were exposed to 0.40 μg/mL of tetra methylthiuram disulphide (TMTD). Doubling times and ATP, ADP and AMP levels were followed on control and treated populations at various cell densities during the exponential growth phase. For the control populations, the average generation time was 171.9 min and ATP levels were 2–8×10−14 mol/cell. For the treated populations, growth was reduced and the populations with the highest densities were more sensitive than populations treated with TMTD at the early stage of the exponential growth phase. For treated populations, ATP and ADP concentrations showed significant differences compared to the control populations. Mechanisms compensating the action of the TMTD have been observed with treated populations at 25,000 cells/mL density. Unfortunately AMP levels were unstable within control populations showing no differences with those from treated populations. ©1999 John Wiley & Sons, Inc. Environ Toxicol 14: 409–413, 1999

Effects of tetramethylthiuram disulphide (thiram) on adenine nucleotide (ATP, ADP, AMP) levels in the ciliateTetrahymena pyriformis

Axenic cultures of Tetrahymena pyriformis GL populations were exposed to 0.40 μg/mL of tetra methylthiuram disulphide (TMTD). Doubling times and ATP, ADP and AMP levels were followed on control and treated populations at various cell densities during the exponential growth phase. For the control populations, the average generation time was 171.9 min and ATP levels were 2–8×10−14 mol/cell. For the treated populations, growth was reduced and the populations with the highest densities were more sensitive than populations treated with TMTD at the early stage of the exponential growth phase. For treated populations, ATP and ADP concentrations showed significant differences compared to the control populations. Mechanisms compensating the action of the TMTD have been observed with treated populations at 25,000 cells/mL density. Unfortunately AMP levels were unstable within control populations showing no differences with those from treated populations. ©1999 John Wiley & Sons, Inc. Environ Toxicol 14: 409–413, 1999