Survival ofL Research Articles

Effects of neem (Azadirachta indica) extracts onLeptocorisa chinensis under choice and no-choice conditions

Two extracts from neem (Azadirachta indica A. Juss. (Meliaceae)) seeds, azadirachtin and oil, and a mixture of neem oil and abamectin, were tested on second-instar nymphs of the rice bugLeptocorisa chinensis (Dallas) (Hemiptera: Alydidae). To clarify the effect of spraying coverage on bioefficacy of test materials, experiments were conducted under choice and no-choice conditions in field cages. In a choice test, treatment with the mixture of neem oil and abamectin was most effective in reducing the survival ofL. chinensis, followed by azadirachtin at 60 ppm, 30 ppm and 3% neem oil, whereas all treatments except neem oil caused 100% mortality within 3 weeks in a no-choice test. When second-instar nymphs had choices of treated and untreated plants within a treatment, no differences in yield and sum of dead and stained grains were found between those two choices, indicating that nymphs neither caused significant reduction in yield nor reduced the quality of untreated plants. Regardless of treatment, the difference in overall yield between treated and untreated plants under choice conditions was not statistically significant (P>0.05). Our results indicate that neem-based formulations, used alone or in combination with abamectin, have the potential to be integrated into the existing programs to control the rice bug.

Identification of sigma factor sigma B-controlled genes and their impact on acid stress, high hydrostatic pressure, and freeze survival in Listeria monocytogenes EGD-e.

The gene encoding the alternative sigma factor sigma(B) in Listeria monocytogenes is induced upon exposure of cells to several stresses. In this study, we investigated the impact of a sigB null mutation on the survival of L. monocytogenes EGD-e at low pH, during high-hydrostatic-pressure treatment, and during freezing. The survival of Delta sigB mutant exponential-phase cells at pH 2.5 was 10,000-fold lower than the survival of EGD-e wild-type cells. Moreover, the Delta sigB mutant failed to show an acid tolerance response. Upon preexposure for 1 h to pH 4.5, the survival at pH 2.5 was 100,000-fold lower for the Delta sigB mutant than for the wild type. The glutamate decarboxylase (GAD) acid resistance system is important in survival and adaptation of L. monocytogenes in acidic conditions. The sigma(B) dependence of the gad genes (gadA, gadB, gadC, gadD, and gadE) was analyzed in silico. Putative sigma(B)-dependent promoter sites were found upstream of the gadCB operon (encoding a glutamate/gamma-aminobutyrate antiporter and a glutamate decarboxylase, respectively) and the lmo2434 gene (gadD, encoding a putative glutamate decarboxylase). Reverse transcriptase PCR revealed that expression of the gadCB operon and expression of gadD are indeed sigma(B) dependent. In addition, a proteomics approach was used to analyze the protein expression profiles upon acid exposure. Although the GAD proteins were not recovered, nine proteins accumulated in the wild type but not in the Delta sigB strain. These proteins included Pfk, GalE, ClpP, and Lmo1580. Exposure to pH 4.5, in order to preload cells with active sigma(B) and consequently with sigma (B)-dependent general stress proteins, also provided considerable protection against high-hydrostatic-pressure treatment and freezing. The combined data argue that the expression of sigma(B)-dependent genes provides L. monocytogenes with nonspecific multiple-stress resistance that may be relevant for survival in the natural environment as well as during food processing.

The RNA-binding protein Hfq of Listeria monocytogenes: role in stress tolerance and virulence.

In gram-negative bacteria, the RNA-binding protein Hfq has emerged as an important regulatory factor in a variety of physiological processes, including stress resistance and virulence. In Escherichia coli, Hfq modulates the stability or the translation of mRNAs and interacts with numerous small regulatory RNAs. Here, we studied the role of Hfq in the stress tolerance and virulence of the gram-positive food-borne human pathogen Listeria monocytogenes. We present evidence that Hfq is involved in the ability of L. monocytogenes to tolerate osmotic and ethanol stress and contributes to long-term survival under amino acid-limiting conditions. However, Hfq is not required for resistance to acid and oxidative stress. Transcription of hfq is induced under various stress conditions, including osmotic and ethanol stress and at the entry into the stationary growth phase, thus supporting the view that Hfq is important for the growth and survival of L. monocytogenes in harsh environments. The stress-inducible transcription of hfq depends on the alternative sigma factor sigmaB, which controls the expression of numerous stress- and virulence-associated genes in L. monocytogenes. Infection studies showed that Hfq contributes to pathogenesis in mice, yet plays no role in the infection of cultured cell lines. This study provides, for the first time, information on the role of Hfq in the stress tolerance and virulence of a gram-positive pathogen.

Role of Listeria monocytogenes sigma(B) in survival of lethal acidic conditions and in the acquired acid tolerance response.

The food-borne pathogen Listeria monocytogenes can acquire enhanced resistance to lethal acid conditions through multiple mechanisms. We investigated contributions of the stress-responsive alternative sigma factor, sigma(B), which is encoded by sigB, to growth phase-dependent acid resistance (AR) and to the adaptive acid tolerance response in L. monocytogenes. At various points throughout growth, we compared the relative survival of L. monocytogenes wild-type and DeltasigB strains that had been exposed to either brain heart infusion (pH 2.5) or synthetic gastric fluid (pH 2.5) with and without prior acid adaptation. Under these conditions, survival of the DeltasigB strain was consistently lower than that of the wild-type strain throughout all phases of growth, ranging from 4 orders of magnitude less in mid-log phase to 2 orders of magnitude less in stationary phase. Survival of both DeltasigB and wild-type L. monocytogenes strains increased by 6 orders of magnitude upon entry into stationary phase, demonstrating that the L. monocytogenes growth phase-dependent AR mechanism is sigma(B) independent. sigma(B)-mediated contributions to acquired acid tolerance appear to be greatest in early logarithmic growth. Loss of a functional sigma(B) reduced the survival of L. monocytogenes at pH 2.5 to a greater extent in the presence of organic acid (100 mM acetic acid) than in the presence of inorganic acid alone (HCl), suggesting that L. monocytogenes protection against organic and inorganic acid may be mediated through different mechanisms. sigma(B) does not appear to contribute to pH(i) homeostasis through regulation of net proton movement across the cell membrane or by regulation of pH(i) buffering by the GAD system under the conditions examined in this study. In summary, a functional sigma(B) protein is necessary for full resistance of L. monocytogenes to lethal acid treatments.

Effect of acid adaptation on the fate of Listeria monocytogenes in THP-1 human macrophages activated by gamma interferon.

In Listeria monocytogenes the acid tolerance response (ATR) takes place through a programmed molecular response which ensures cell survival under unfavorable conditions. Much evidence links ATR with virulence, but the molecular determinants involved in the reactivity to low pHs and the behavior of acid-exposed bacteria within host cells are still poorly understood. We have investigated the effect of acid adaptation on the fate of L. monocytogenes in human macrophages. Expression of genes encoding determinants for cell invasion and intracellular survival was tested for acid-exposed bacteria, and invasive behavior in the human myelomonocytic cell line THP-1 activated with gamma interferon was assessed. Functional approaches demonstrated that preexposure to an acidic pH enhances the survival of L. monocytogenes in activated human macrophages and that this effect is associated with an altered pattern of expression of genes involved in acid resistance and cell invasion. Significantly decreased transcription of the plcA gene, encoding a phospholipase C involved in vacuolar escape and cell-to-cell spread, was observed in acid-adapted bacteria. This effect was due to a reduction in the quantity of the bicistronic plcA-prfA transcript, concomitant with an increase in the level(s) of the monocistronic prfA mRNA(s). The transcriptional shift from distal to proximal prfA promoters resulted in equal levels of the prfA transcript (and, as a consequence, of the inlA, hly, and actA transcripts) under neutral and acidic conditions. In contrast, the sodC and gad genes, encoding a cytoplasmic superoxide dismutase and the glutamate-based acid resistance system, respectively, were positively regulated at a low pH. Morphological approaches confirmed the increased intracellular survival and growth of acid-adapted L. monocytogenes cells both in vacuoles and in the cytoplasm of interferon gamma-activated THP-1 macrophages. Our data indicate that preexposure to a low pH has a positive impact on subsequent challenge of L. monocytogenes with macrophagic cells.

Microhabitat availability and seedling recruitment ofLobelia urens: a rare plant species at its geographical limit

AbstractIn Britain,Lobelia urens(L.) (the heath lobelia) occurs in rough grassland, is rare and only found in southern England, where it is at the northern limit of its range. Emergence and survival ofL. urenswas investigated at six locations in two geographically distinct sites experiencing spring, autumn or no grazing in two consecutive years. Four factors were evaluated qualitatively, as a means of characterizing microhabitats for germination and survival: all permutations of higher plant cover, bryophytes, plant litter and surface depressions. The potential effect of adjacent plants on recruitment was also assessed using the nearest neighbour distance (NND). Grazing created depressions, removed plant litter and increased the proportion of sites with higher plant cover. It also resulted in a more open sward with higher NNDs. None of these changes stimulated recruitment. Instead, small increases in the frequency of some rare or very rare microhabitat types were vital in making grazed rough grassland more suitable for emergence. Reduced litter loads and a greater quantity of moss were two key responses. Overall, survival ofL. urenswas less than 1% and was particularly favoured by moss and an increase in NNDs. Shading from higher plants, with or without plant litter, decreased emergence, but the precise role of litter was complex and most probably related to its quantity. In an experimental seed bed, only empty depressions favoured emergence. The microhabitat relationships ofL. urenswere unusually consistent among locations and consecutive years.L. urensrequires high soil surface temperatures but also adequate water for large scale recruitment, and such conditions are encouraged by grazing. The particular problems of experimental design and statistical analysis of data from recruitment experiments are also discussed.

Induction of stress proteins inLeuconostoc oenos to perform direct inoculation of wine

The enhancement or induction of the protein synthesis was clearly observed in cells ofL. oenos labeled with35S for five proteins during heat shock at 42°C and acid shock at pH 3. Furthermore, no stress protein was induced after exposure ofL. oenos to ethanol shock 10% (v/v). Moreover, survival ofL. oenos in wine and ability to perform alolactic fermentation was improved after direct inoculation when cells were pretreated at 42°C.

Survival ofLegionella pneumophila in the aquatic environment

Survival ofLegionella pneumophila SG 1 in seawater and river water was assessed using plate counts on buffered charcoal yeast extract agar amended with α-ketoglutarate (BCYEα) and [(3)H]thymidine-labeling. The [(3)H]thymidine-labeling method for assessing survival ofL. pneumophila in aquatic environments was compared with viable counts, direct fluorescent microscopy (DFA), and acridine orange direct counts (AODC). Protozoa were isolated from the samples employed in the study and identified by characteristic trophozite and cyst morphology. Selective filtration employing 2.0 μm Nucleopore filters was used to determine the effect of grazing on survival ofL. pneumophila in seawater and river water.Legionella viability as measured by plate counts (CFU/ml), declined to a greater extent than cell lysis, assessed by thymidine, DFA, and AODC counts, suggesting thatL. pneumophila survives in aquatic habitats to a greater extent than revealed through culturable counts.

A perforated gastrovascular cavity in the symbiotic deep-water coralLeptoseris fragilis: A new strategy to optimize heterotrophic nutrition

The organization of the zooxanthellate scleractinian coralLeptoseris fragilis was studied. The architecture of the corallite and the histology of the polyparium were analysed for adaptations that enable efficient capture and retention of suspended particles which would increase energy supply. The data indicate that the gastrovascular system ofL. fragilis is not a blind but a flowthrough system. Water entering the coelenteron through the mouth leaves the body not only through the mouth but also through microscopic pores (≂ 1–2 μm) which are located near the crests of the sclerosepta in the oral epithelia. Irrigation is achieved by flagellar and probably also by muscular activity. This type of filtration enablesL. fragilis, which lacks tentacles, to utilize suspended organic material including bacteria. The supposed suspension feeding in combination with effective photoadaptations (presented in former communications) seems to be the basis for the survival ofL. fragilis in an extreme habitat (between-95 and-145 m) and for its, successful competion with other scleractinian species provided with larger catching surfaces, and with other invertebrates depending on filter feeding.

Host habitat finding and host selection of theDrosophila parasitoidLeptopilina australis (Hymenoptera, Eucoilidae), with a comparison of the niches of EuropeanLeptopilina species.

Decaying petioles of giant hogweed,Heracleum mantegazzianum Sommier & Levier, are used as a breeding site by six species ofDrosophila and the drosophilidScaptomyza pallida. The most numerous parasitoid species associated with this community isLeptopilina australis. BecauseL. australis was previously unknown in western Europe, we present the characters to distinguish it form its close relativeL. clavipes. Experiments on host species selection and survival ofL. australis showed that this parasitoid mainly usesD. limbata as host. Olfactometer experiments showed thatL. australis is attracted by the odour of decaying hogweed stalks, especially when these contain larvae ofD. limbata. L. australis is also strongly attracted by the odour of stinkhorns, a habitat in which it has never been found in nature.D. phalerata is the dominant fly species in stinkhorns, and is not a host ofL. australis. We offer a possible functional explanation for this unexpected habitat choice, by showing thatD. transversa andD. kuntzei, both species found to breed in fungi, are also suitable hosts forL. australis. We also discuss habitat choice with regard to a proposed phylogeny of theLeptopilina species in temperate Europe. Finally, we discuss niche overlap ofL. australis with the otherLeptopilina species.

The effect of the nanoflagellateTetraselmis suecica on the growth and survival of grunion,Leuresthes tenuis, larvae

The nanoflagellateTetraselmis suecica was tested both as the sole food source and as a diet complement toArtemia nauplii for grunion,Leuresthes tenuis, larvae. A total of 4800 grunion larvae, obtained through artificial insemination and incubation, were cultivated under laboratory conditions. Growth and survival rates were registered for 14 days in two experimental series. In the first series the nanoflagellateT. suecica was offered as the sole food source at five different concentrations. Survival and growth increased in agreement with the increase inT. suecica concentration. In the second series,Artemia nauplii were offered at six concentration levels. This series was divided into two groups: the nanoflagellateT. suecica was added to one group at a concentration of 5000 cells ml−1; the other group was maintained without nanoflagellates. In this series, survival and growth were directly related to nauplii concentration, but significant effects of the nanoflagellates were evident only in relation to the survival; the greatest difference (58% without nanoflagellates vs. 69% with nanoflagellates) was observed at anArtemia concentration of 1000 nauplii 1−1. The mechanism responsible for increased survival ofL. tenuis larvae in presence of phytoplankton is unclear.

Obligate mixotrophy inLaboea strobila, a ciliate which retains chloroplasts

The planktonic ciliateLaboea strobila Lohmann sequesters photosynthetically functional chloroplasts derived from ingested algae. The chloroplasts lie free in the cytoplasm and are most abundant just under the pellicle of the ciliate. The maximum rate of photosynthesis (Pmax) was 925 pg C ciliate-1h-1 (3.7 pg C pg chl.a-1h-1). At saturating irradiance, the amount of carbon fixed h-1 equaled 12.6% of the body carbon of the ciliate. To grow,L. strobila requires both light and algal food. In the absence of food, survival ofL. strobila is significantly longer in the light than in the dark. Based on ingestion rate and photosynthetic rate, we calculate that photosynthesis can make an important contribution to this ciliate's carbon budget even when algal food is plentiful.