Streptomycin Induced Stress Response in Salmonella enterica Serovar Typhimurium Shows Distinct Colony Scatter Signature

We investigated the streptomycin-induced stress response in Salmonella enterica serovars with a laser optical sensor, BARDOT (bacterial rapid detection using optical scattering technology). Initially, the top 20 S. enterica serovars were screened for their response to streptomycin at 100 μg/mL. All, but four S. enterica serovars were resistant to streptomycin. The MIC of streptomycin-sensitive serovars (Enteritidis, Muenchen, Mississippi, and Schwarzengrund) varied from 12.5 to 50 μg/mL, while streptomycin-resistant serovar (Typhimurium) from 125–250 μg/mL. Two streptomycin-sensitive serovars (Enteritidis and Mississippi) were grown on brain heart infusion (BHI) agar plates containing sub-inhibitory concentration of streptomycin (1.25–5 μg/mL) and a streptomycin-resistant serovar (Typhimurium) was grown on BHI containing 25–50 μg/mL of streptomycin and the colonies (1.2 ± 0.1 mm diameter) were scanned using BARDOT. Data show substantial qualitative and quantitative differences in the colony scatter patterns of Salmonella grown in the presence of streptomycin than the colonies grown in absence of antibiotic. Mass-spectrometry identified overexpression of chaperonin GroEL, which possibly contributed to the observed differences in the colony scatter patterns. Quantitative RT-PCR and immunoassay confirmed streptomycin-induced GroEL expression while, aminoglycoside adenylyltransferase (aadA), aminoglycoside efflux pump (aep), multidrug resistance subunit acrA, and ribosomal protein S12 (rpsL), involved in streptomycin resistance, were unaltered. The study highlights suitability of the BARDOT as a non-invasive, label-free tool for investigating stress response in Salmonella in conjunction with the molecular and immunoassay methods.     

Plant-Adapted Escherichia coli Show Increased Lettuce Colonizing Ability,Resistance to Oxidative Stress and Chemotactic Response

Background

Escherichia coli is a widespread gut commensal and often a versatile pathogen of public health concern. E. coli are also frequently found in different environments and/or alternative secondary hosts, such as plant tissues. The lifestyle of E. coli in plants is poorly understood and has potential implications for food safety.

Methods/Principal Findings

This work shows that a human commensal strain of E. coli K12 readily colonizes lettuce seedlings and produces large microcolony-like cell aggregates in leaves, especially in young leaves, in proximity to the vascular tissue. Our observations strongly suggest that those cell aggregates arise from multiplication of single bacterial cells that reach those spots. We showed that E. coli isolated from colonized leaves progressively colonize lettuce seedlings to higher titers, suggesting a fast adaptation process. E. coli cells isolated from leaves presented a dramatic rise in tolerance to oxidative stress and became more chemotactic responsive towards lettuce leaf extracts. Mutant strains impaired in their chemotactic response were less efficient lettuce colonizers than the chemotactic isogenic strain. However, acclimation to oxidative stress and/or minimal medium alone failed to prime E. coli cells for enhanced lettuce colonization efficiency.

Conclusion/Significance

These findings help to understand the physiological adaptation during the alternative lifestyle of E. coli in/on plant tissues.     

Preadaptation to Cold Stress in Salmonella enterica Serovar Typhimurium Increases Survival during Subsequent Acid Stress Exposure

Salmonella is an important cause of bacterial food-borne gastroenteritis. Salmonella encounters multiple abiotic stresses during pathogen elimination methods used in food processing, and these stresses may influence its subsequent survivability within the host or in the environment. Upon ingestion, Salmonella is exposed to gastrointestinal acidity, a first line of the host innate defense system. This study tested the hypothesis that abiotic stresses encountered during food processing alter the metabolic mechanisms in Salmonella that enable survival and persistence during subsequent exposure to the host gastrointestinal acidic environment. Out of the four different abiotic stresses tested, viz., cold, peroxide, osmotic, and acid, preadaptation of the log-phase culture to cold stress (5°C for 5 h) significantly enhanced survival during subsequent acid stress (pH 4.0 for 90 min). The gene expression profile of Salmonella preadapted to cold stress revealed induction of multiple genes associated with amino acid metabolism, oxidative stress, and DNA repair, while only a few of the genes in the above-mentioned stress response and repair pathways were induced upon exposure to acid stress alone. Preadaptation to cold stress decreased the NAD⁺/NADH ratio and hydroxyl (OH·) radical formation compared with those achieved with the exposure to acid stress alone, indicating alteration of aerobic respiration and the oxidative state of the bacteria. The results from this study suggest that preadaptation to cold stress rescues Salmonella from the deleterious effect of subsequent acid stress exposure by induction of genes involved in stress response and repair pathways, by modification of aerobic respiration, and by redox modulation.     

Oxidative Stress and Heat Shock Protein Induction in Human Cells

Agents which induce heat shock protein synthesis in cultured monolayers of Hela cells such as hyperthermia, ethanol and sodium arsenite can also cause increases in the levels of lipid peroxidation as determined by the formation of TBA-products. The heat induced increases may be diminished by addition to the medium of mannitol or EGTA. These compounds are known to depress heat shock protein synthesis.

Following hyperthermia there is also a decrease in protein synthesis. In vitro studies indicate possible damage to ribosomes, and since the heat induced loss of protein synthetic capacity can be increased by superoxide dismutase inhibitors, and prevented by mannitol, such effects may be linked to the increases observed in lipid peroxidation. It is suggested that a connection exists between lipid peroxidation and heat shock protein gene activation.     

Oxidative Stress and Heat Shock Protein Induction in Human Cells

Agents which induce heat shock protein synthesis in cultured monolayers of Hela cells such as hyperthermia, ethanol and sodium arsenite can also cause increases in the levels of lipid peroxidation as determined by the formation of TBA-products. The heat induced increases may be diminished by addition to the medium of mannitol or EGTA. These compounds are known to depress heat shock protein synthesis.

Following hyperthermia there is also a decrease in protein synthesis. In vitro studies indicate possible damage to ribosomes, and since the heat induced loss of protein synthetic capacity can be increased by superoxide dismutase inhibitors, and prevented by mannitol, such effects may be linked to the increases observed in lipid peroxidation. It is suggested that a connection exists between lipid peroxidation and heat shock protein gene activation.     

Response of Two Heat Shock Genes to Selection for Knockdown Heat Resistance in Drosophila Melanogaster

To identify genes involved in stress resistance and heat hardening, replicate lines of Drosophila melanogaster were selected for increased resistance to knockdown by a 39° heat stress. Two selective regimes were used, one with and one without prior hardening. Mean knockdown times were increased from ~5 min to >20 min after 18 generations. Initial realized heritabilities were as high as 10% for lines selected without hardening, and crosses between lines indicated simple additive gene effects for the selected phenotypes. To survey allelic variation and correlated selection responses in two candidate stress genes, hsr-omega and hsp68, we applied denaturing gradient gel electrophoresis to amplified DNA sequences from small regions of these genes. After eight generations of selection, allele frequencies at both loci showed correlated responses for selection following hardening, but not without hardening. The hardening process itself was associated with a hsp68 frequency change in the opposite direction to that associated with selection that followed hardening. These stress loci are closely linked on chromosome III, and the hardening selection established a disequilibrium, suggesting an epistatic effect on resistance. The data indicate that molecular variation in both hsr-omega and hsp68 contribute to natural heritable variation for hardened heat resistance.     

工业微生物的酸胁迫响应及其抵御策略

有机酸的积累在工业发酵过程中是一个较为普遍的现象,会导致发酵体系pH的降低,进而引起酸胁迫,限制细胞生长及目标产物的积累。针对这一问题,本文阐述了工业微生物应对酸胁迫所发生的生理变化,提出了微生物在酸性环境中可能的3种自我调节机制,概括总结了目前提高微生物酸耐受性的策略及各自的优缺点,指出了未来可能的发展方向,以期为提高工业微生物的酸耐受性提供思路。     

Heat Shock Response and Organ Preservation: Models of Stress Conditioning

No Abstract Available     

Response of Higher Plants to Heat Shock

When sorghum seedlings were rapidly shifted from the cultural temperature of 30℃ to 40℃ and 45℃, a set of abnormal proteins, generally referred to as heat shock proteins were induced. They are a group of high molecular weight proteins (about 66–117 kD), a few intermediate molecular weight proteins (33–66kD) and a low molecular weight protein of 18 kD. At the same time, the synthesis of normal proteins was relatively depressed. The res ponse of the shoot tissues of sorghum seedings to heat shock is similar to that of the root tissues, but there are some differences in more detail between the two tissues. The synthesis of heat shock proteins in sorghum seedlings was rapid. After one-hour exposure at 45℃ their synthesis in the roots was detectable. Maximum induction took place in the second hour of exposure, thereafter their synthesis began to decline markedly. Finally, there appear to be some proteins whose synthesis was not supressed during heat shock, It is not yet known why the synthesis of these proteins is so stable.     

Effect of Heat Shock on the mRNA-Directed Disease Resistance Response of Peas

Pea tissue `heat shocked' for 2 hours at 40°C accumulates mRNAs that code for a series of new proteins called heat shock proteins. A different messenger RNA population, which codes for a high level of 20 or more `resistance proteins,' accumulates in pea tissue as it resists plant pathogenic fungi. Heat shock treatment applied prior to fungal inoculation prevents the high level accumulation of messenger RNA coding for the 20 resistance proteins and blocks disease resistance. If the resistance response is initiated before the heat shock treatment or after heat shock recovery, messenger RNA accumulates for the resistance proteins and disease resistance develops.     

Changes in Ultrastructure and Fourier Transform Infrared Spectrum of Salmonella enterica Serovar Typhimurium Cells after Exposure to Stress Conditions

The effect of exposure to acid (pH 2.5), alkaline (pH 11.0), heat (55°C), and oxidative (40 mM H₂O₂) lethal conditions on the ultrastructure and global chemical composition of Salmonella enterica serovar Typhimurium CECT 443 cells was studied using transmission electron microscopy and Fourier transform infrared spectroscopy (FT-IR) combined with multivariate statistical methods (hierarchical cluster analysis and factor analysis). Infrared spectra exhibited marked differences in the five spectral regions for all conditions tested compared to those of nontreated control cells, which suggests the existence of a complex bacterial stress response in which modifications in a wide variety of cellular compounds are involved. The visible spectral changes observed in all of the spectral regions, together with ultrastructural changes observed by transmission electron microscopy and data obtained from membrane integrity tests, indicate the existence of membrane damage or alterations in membrane composition after heat, acid, alkaline, and oxidative treatments. Results obtained in this study indicate the potential of FT-IR spectroscopy to discriminate between intact and injured bacterial cells and between treatment technologies, and they show the adequacy of this technique to study the molecular aspects of bacterial stress response.Salmonella spp. are an important cause of bacterial food-borne disease all over the world, causing a diversity of illnesses that include typhoid fever, gastroenteritis, and septicemia (11). According to epidemiological data from the European Union, a total of 131,468 laboratory-confirmed salmonellosis cases were reported in 2008, with two serovars, Salmonella enterica serovar Typhimurium and Salmonella enterica serovar Enteritidis, being responsible for 79.9% of all cases (13). The detection and identification of pathogens in foods are a basic cornerstone of food safety, because they make it possible to identify sources of contamination, provide data on the evaluation of risk reduction measures, and identify the food chain operations, processes, batches, or products representing a threat to public health. Furthermore, they also are fundamental in the epidemiological investigation of food-borne diseases. The presence of stress-injured bacterial cells in foods represents a challenge to those involved in food quality assurance, as routine microbiological procedures may yield negative results for sublethally injured cells. Thus, food could be presumed to be safe and free from pathogenic cells but during storage become dangerous due to the recovery and growth of previously injured cells.Given the fact that bacterial cells react to the different environmental stress conditions by inducing structural and physiological changes, Fourier transform infrared (FT-IR) spectroscopy, which reflects the biochemical composition of the cellular constituents of bacteria that include water, fatty acids, proteins, polysaccharides, and nucleic acids (26), should be able to monitor the changes occurring in bacterial cells in response to several food-related stress conditions. The potential of this methodology to detect and differentiate sublethally heat-injured and dead Listeria monocytogenes and S. Typhimurium cells and to discriminate between diverse heat treatment intensities has been highlighted (2, 20). FT-IR spectroscopy also has been successfully applied to the identification and classification of bacteria such as Acinetobacter (35), Brucella (23), Campylobacter (24, 25), Escherichia coli (1), Lactobacillus (10, 27), Listeria (28, 29), Salmonella (9, 17), Staphylococcus (8, 19), and Yersinia (18).The main aim of this study was to assess ultrastructural modifications and infrared (IR) spectral changes at different degrees of stress exposure and to discriminate between injured and noninjured S. Typhimurium cells after their exposure to heat, acid, alkaline, and oxidative lethal conditions. Results obtained also could help us improve our knowledge of S. Typhimurium cell damage and strategies of response to these adverse conditions.     

Response of Mutant Sunflower Lines to Heat Shock

The effects of heat shock (HS) (40°C for 1 h) on the level of malondialdehyde (MDA), the terminal product of lipid peroxidation, superoxide dismutase (SOD) activity, catalase activity, and total peroxidase activity (TPA) were studied in root meristems and chloroplasts of several sunflower (Helianthus annuusL.) lines that carried nuclear or plastome chlorophyll mutations. HS either lowered or did not affect the MDA level in the root meristem and in the chloroplasts from the first true leaf, as compared to the untreated plants. In both treatments, the root and leaf enzyme activities varied in the sunflower lines. In the root meristem, catalase was the most sensitive to HS, whereas, in the chloroplasts from HS-treated sunflower lines, HS activated either TPA or SOD.