Sublethal Heat Research Articles

Activation of AMP-activated protein kinase in response to temperature elevation shows seasonal variation in the zebra mussel, Dreissena polymorpha

Global climate change is affecting ectothermic species, and a variety of studies are needed on thermal tolerances, especially from cellular and physiological perspectives. This study utilized AMP-activated protein kinase (AMPK), a key regulator of cellular energy levels, to examine the effects of high water temperatures on zebra mussel (Dreissena polymorpha) physiology. During heating, AMPK activity increased as water temperature increased to a point, and maximum AMPK activity was detected at high, but sublethal, water temperatures. This pattern varied with season, suggesting that cellular mechanisms of seasonal thermal acclimatization affect basic metabolic processes during sublethal heat stress. There was a greater seasonal variation in the water temperature at which maximum AMPK activity was measured than in lethal water temperature. Furthermore, baseline AMPK activity varied significantly across seasons, most likely reflecting altered metabolic states during times of growth and reproduction. In addition, when summer-collected mussels were lab-acclimated to winter and spring water temperatures, patterns of heat stress mirrored those of field-collected animals. These data suggest that water temperature is the main driver of the seasonal variation in physiology. This study concluded that AMPK activity, which reflects changes in energy supply and demand during heat stress, can serve as a sensitive and early indicator of temperature stress in mussels.

Application of heat stress in situ demonstrates a protective role of irradiation on photosynthetic performance in alpine plants.

The impact of sublethal heat on photosynthetic performance, photosynthetic pigments and free radical scavenging activity was examined in three high mountain species, Rhododendron ferrugineum, Senecio incanus and Ranunculus glacialis using controlled in situ applications of heat stress, both in darkness and under natural solar irradiation. Heat treatments applied in the dark reversibly reduced photosynthetic performance and the maximum quantum efficiency of photosystem II (Fv/Fm), which remained impeded for several days when plants were exposed to natural light conditions subsequently to the heat treatment. In contrast, plants exposed to heat stress under natural irradiation were able to tolerate and recover from heat stress more readily. The critical temperature threshold for chlorophyll fluorescence was higher under illumination (Tc′) than in the dark (Tc). Heat stress caused a significant de-epoxidation of the xanthophyll cycle pigments both in the light and in the dark conditions. Total free radical scavenging activity was highest when heat stress was applied in the dark. This study demonstrates that, in the European Alps, heat waves can temporarily have a negative impact on photosynthesis and, importantly, that results obtained from experiments performed in darkness and/or on detached plant material may not reliably predict the impact of heat stress under field conditions.

Evaluation of Salmonella Thermal Inactivation Model Validity for Slow Cooking of Whole-Muscle Meat Roasts in a Pilot-Scale Oven

Sublethal heating can increase subsequent thermal resistance of bacteria, which may compromise the validity of thermal process validations for slow-roasted meats. Therefore, this research evaluated the accuracy of a traditional log-linear inactivation model, developed via prior laboratory-scale isothermal tests, and a novel path-dependent model accounting for sublethal injury, applied to pilot-scale slow cooking of whole-muscle roasts. Irradiated turkey breasts, beef rounds, and pork loins were inoculated with an eight-serovar Salmonella cocktail via vacuum tumble marination in a salt-phosphate marinade. The resulting initial Salmonella population in the geometric center (core) was 7.0, 6.3, and 6.3 log CFU/g for turkey, beef, and pork, respectively. Seven different cooking schedules representing industry practices were evaluated in a pilot-scale, moist-air convection oven. Core temperatures recorded during cooking were used to calculate lethality real-time via the log-linear model. The path-dependent model reduced the bias (mean residual) and root mean square error by 4.24 and 4.60 log CFU/g respectively, in turkey; however, the new model did not reduce the prediction error in beef or pork. Overall, results demonstrated that slow-cooked roasts, processed to a computed lethality at or near that required by the regulatory performance standards, as calculated with a state-dependent model, may be underprocessed.

Environmental and biological factors influencing the UV-C resistance of Listeria monocytogenes

In this investigation, the effect of microbiological factors (strain, growth phase, exposition to sublethal stresses, and photorepair ability), treatment medium characteristics (pH, water activity, and absorption coefficient), and processing parameters (dose and temperature) on the UV resistance of Listeria monocytogenes was studied. The dose to inactivate 99.99% of the initial population of the five strains tested ranged from 21.84 J/mL (STCC 5672) to 14.66 J/mL (STCC 4031). The UV inactivation of the most resistant strain did not change in different growth phases and after exposure to sublethal heat, acid, basic, and oxidative shocks. The pH and water activity of the treatment medium did not affect the UV resistance of L. monocytogenes, whereas the inactivation rate decreased exponentially with the absorption coefficient. The lethal effect of UV radiation increased synergistically with temperature between 50 and 60 °C (UV-H treatment). A UV-H treatment of 27.10 J/mL at 55 °C reached 2.99 and 3.69 Log10 inactivation cycles of L. monocytogenes in orange juice and vegetable broth, and more than 5 Log10 cycles in apple juice and chicken broth. This synergistic effect opens the possibility to design UV combined processes for the pasteurization of liquid foods with high absorptivity.

Low, Medium, and High Heat Tolerant Strains of Listeria monocytogenes and Increased Heat Stress Resistance after Exposure to Sublethal Heat

A group of 37 strains representing all 13 serotypes of Listeria monocytogenes with an initial cell density of 10(7) CFU/ml were analyzed for their heat tolerance at 60°C for 10 min. These L. monocytogenes strains were categorized into three heat tolerance groups: low (<2 log CFU/ml survival), medium (2 to 4 log CFU/ml survival), and high (4 to 6 log CFU/ml survival). Serotype 1/2a strains had relatively low heat tolerance; seven of the eight tested strains were classified as low heat tolerant. Of the two serotype 1/2b strains tested, one was very heat sensitive (not detectable) and the other was very heat resistant (5.4 log CFU/ml survival). Among the 16 serotype 4b strains, survival ranged from not detectable to 4 log CFU/ml. When one L. monocytogenes strain from each heat tolerance group was subjected to sublethal heat stress at 48°C for 30 or 60 min, the survival of heat-stressed cells at 60°C for 10 min increased by 5 log CFU/ml (D60°C-values nearly doubled) compared with the nonstressed control cells. Sublethal heat stress at 48°C for 60 or 90 min increased the lag phase of L. monocytogenes in tryptic soy broth supplemented with 0.6% yeast extract at room temperature by 3 to 5 h compared with nonstressed control cells. The heat stress adaptation in L. monocytogenes was reversed after 2 h at room temperature but was maintained for up to 24 h at 4°C. Our results indicate a high diversity in heat tolerance among strains of L. monocytogenes, and once acquired this heat stress adaptation persists after cooling, which should be taken into account while conducting risk analyses for this pathogen.

Sub-lethal heat stress causes apoptosis in an Antarctic fish that lacks an inducible heat shock response

The endemic fish fauna of the Southern Ocean are cold-adapted stenotherms and are acutely sensitive to elevated temperature. Many of these species lack a heat shock response and cannot increase the production of heat shock proteins in their tissues. However, some species retain the ability to induce other stress-responsive genes, some of which are involved in cell cycle arrest and apoptosis. Here, the effect of heat on cell cycle stage and its ability to induce apoptosis were tested in thermally stressed hepatocytes from a common Antarctic fish species from McMurdo Sound in the Ross Sea. Levels of proliferating cell nuclear antigen were also measured as a marker of progression through the cell cycle. The results of these studies demonstrate that even sub-lethal heat stress can have deleterious impacts at the cellular level on these environmentally sensitive species.

Membrane fluidity and temperature sensing are coupled via circuitry comprised of Ole1, Rsp5, and Hsf1 in Candida albicans.

Temperature is a ubiquitous environmental variable which can profoundly influence the physiology of living cells as it changes over time and space. When yeast cells are exposed to a sublethal heat shock, normal metabolic functions become repressed and the heat shock transcription factor Hsf1 is activated, inducing heat shock proteins (HSPs). Candida albicans, the most prevalent human fungal pathogen, is an opportunistic pathogen that has evolved as a relatively harmless commensal of healthy individuals. Even though C. albicans occupies thermally buffered niches, it has retained the classic heat shock response, activating Hsf1 during slow thermal transitions such as the increases in temperature suffered by febrile patients. However, the mechanism of temperature sensing in fungal pathogens remains enigmatic. A few studies with Saccharomyces cerevisiae suggest that thermal stress is transduced into a cellular signal at the level of the membrane. In this study, we manipulated the fluidity of C. albicans membrane to dissect mechanisms of temperature sensing. We determined that in response to elevated temperature, levels of OLE1, encoding a fatty acid desaturase, decrease. Subsequently, loss of OLE1 triggers expression of FAS2, encoding a fatty acid synthase. Furthermore, depletion of OLE1 prevents full activation of Hsf1, thereby reducing HSP expression in response to heat shock. This reduction in Hsf1 activation is attributable to the E3 ubiquitin ligase Rsp5, which regulates OLE1 expression. To our knowledge, this is the first study to define a molecular link between fatty acid synthesis and the heat shock response in the fungal kingdom.

Evidence for adaptive divergence of thermal responses among Bemisia tabaci populations from tropical Colombia following a recent invasion.

There is an increasing evidence that populations of ectotherms can diverge genetically in response to different climatic conditions, both within their native range and (in the case of invasive species) in their new range. Here, we test for such divergence in invasive whitefly Bemisia tabaci populations in tropical Colombia, by considering heritable variation within and between populations in survival and fecundity under temperature stress, and by comparing population differences with patterns established from putatively neutral microsatellite markers. We detected significant differences among populations linked to mean temperature (for survival) and temperature variation (for fecundity) in local environments. A QST -FST analysis indicated that phenotypic divergence was often larger than neutral expectations (QST >FST ). Particularly, for survival after a sublethal heat shock, this divergence remained linked to the local mean temperature after controlling for neutral divergence. These findings point to rapid adaptation in invasive whitefly likely to contribute to its success as a pest species. Ongoing evolutionary divergence also provides challenges in predicting the likely impact of Bemisia in invaded regions.

Sublethal heat treatment promotes epithelial-mesenchymal transition and enhances the malignant potential of hepatocellular carcinoma

Sublethal heat treatment promotes epithelial-mesenchymal transition and enhances the malignant potential of hepatocellular carcinoma

Temporal patterns of cardiac performance and genes encoding heat shock proteins and metabolic sensors of an intertidal limpet Cellana toreuma during sublethal heat stress

Intertidal invertebrates develop effective physiological adaptations to cope with the rapidly changing thermal environment in the intertidal zone. In the present study, the temporal patterns of heart rate, protein carbonyl groups, and genes encoding heat shock proteins (hsp70 and hsp90) and metabolic sensors (ampkα, ampkβ and sirt1) were measured to study the effect of sublethal heat stress on the cardiac function, oxidative stress, heat shock response and cellular metabolism of an intertidal limpet Cellana toreuma. All the physiological parameters are sensitive to temperature and duration of heat stress. Spearman correlation analysis revealed that the correlations between heart rate and levels of heat shock proteins mRNA and metabolic sensors mRNA were statistically significant. These results further suggest that cardiac function plays crucial roles in cellular energy metabolism and heat shock responses. The significant increase of protein carbonyl groups at 34°C after 4h exposure indicated that the failure of cardiac function and the increase of anaerobic metabolism partly leads to the increase of protein carbonyl groups. Generally, the physiological responses to heat stress are sensitive to temperature and are energy-consumptive, as indicated by the upregulation of metabolic sensors mRNA. However, the upregulation of heat shock proteins and metabolic sensors at the post-transcriptional level and related functions need to be confirmed in further experiments.

Role for Putative Hepatocellular Carcinoma Stem Cell Subpopulations in Biological Response to Incomplete Thermal Ablation: In Vitro and In Vivo Pilot Study

To investigate the potential role for CD44(+) and CD90(+) hepatocellular carcinoma (HCC) cellular subpopulations in biological response to thermal ablation-induced heat stress. This study was approved by the institutional animal care committee. The N1S1 rat HCC cell line was subjected to sublethal heat stress (45 °C) or control (37 °C) for 10 min, costained with fluorescent-labeled antibodies against CD44, CD90, and 7-AAD after a 48-h recovery and analyzed by flow cytometry to assess the percentage of live CD44(+) and CD90(+) HCC cells (n = 4). Experiments were repeated with pretreatment of N1S1 cells with a dose titration of the dual PI3K-mTOR inhibitor BEZ235 or vehicle control (n = 3). Rats bearing orthotopic N1S1 tumors were subjected to ultrasound-guided partial laser ablation (n = 5) or sham ablation (n = 3), euthanized 24 h after ablation, and liver/tumor analyzed for immunohistochemical staining of CD44 and CD90. Differences between groups were compared with an unpaired t test. Sublethal heat stress induced a significant increase in the relative proportion of live CD44(+) and CD90(+) HCC cells compared to the control group: CD44(+)CD90(-) (5.3-fold; p = 0.0001), CD44(-)CD90(+) (2.4-fold; p = 0.003), and CD44(+)CD90(+) (22.0-fold; p < 0.03). Inhibition of PI3K-mTOR prevented heat stress-induced enrichment of the population of live CD44(+) HCC cells (p < 0.01), but not CD90(+) cells (p > 0.10). Immunohistochemical analysis demonstrated preferential localization of clusters of CD44(+) cells at both the tumor margin and ablation margin. These studies provide experimental evidence supporting a role for HCC cells expressing the putative stem cell marker CD44 in HCC response to heat stress.

Inhibitory effect of enterocin KP in combination with sublethal factors on Escherichia coli O157:H7 or Salmonella Typhimurium in BHI broth and UHT milk

The effects of physical and chemical sublethal treatments on the antibacterial activity of enterocin KP produced by Enterococcus faecalis KP against Escherichia coli O157:H7 and Salmonella Typhimurium were investigated. Enterocin KP was not active against intact cells of E. coli O157:H7 or S. Typhimurium. However, the use of enterocin KP together with ethylenediaminetetraacetic acid (50 mM), sodium tripolyphosphate (50 mM), sublethal heating (60 °C for 10 min), cold shock (-20 °C for 2 h), or acid stress (mixture of 40% lactic acid, 16% propionic acid, 16% acetic acid) in BHI medium decreased the cell number of E. coli O157:H7 by 7.27, 6.28, 3.39, 3.06, 4.20 log and S. Typhimurium by 7.21, 6.20, 3.64, 3.38, 3.98 log cfu/mL, respectively. The combination of enterocin KP with ethylenediaminetetraacetic acid decreased the cell number of E. coli O157:H7 in UHT milk to undetectable level, enterocin KP plus sodium tripolyphosphate or enterocin KP plus sublethal heating caused a reduction by 6.07 and 5.68 log cycles. The results of this study showed that enterocin KP could be applied as a biopreservative to inhibit E. coli O157:H7 and S. Typhimurium in combination with physical and food grade chemical hurdles.

Peripheral blood mononuclear cells: a potential cellular system to understand differential heat shock response across native cattle (Bos indicus), exotic cattle (Bos taurus), and riverine buffaloes (Bubalus bubalis) of India

Circulating leukocytes can be used as an effective model to understand the heat stress response of different cattle types and buffaloes. This investigation aimed to determine the temporal profile of HSPs (HSP40, HSP60, HSP70, and HSP90) expression in circulating peripheral blood mononuclear cells (PBMCs) of Murrah buffaloes, Holstein-Friesian (HF), and Sahiwal cows in response to sublethal heat shock at 42 °C. The viability data indicated HF PBMCs to be the most affected to the heat shock, whereas Sahiwal PBMCs were least affected, indicating its better survivability during the heat stress condition. The qRT-PCR expression data showed significant increase in mRNA expression of the analyzed HSPs genes after heat stimuli to the PBMCs under in vitro condition. In each case, the HSPs were most upregulated at 2 h after the heat stress. Among the HSPs, HSP70 was relatively more expressed followed by HSP60 indicating the action of molecular chaperones to stabilize the native conformation of proteins. However, PBMCs from different cattle types and buffaloes showed difference in the extent of transcriptional response. The level of expression of HSPs throughout the time period of heat stress was highest in buffaloes, followed by HF and Sahiwal cows. The higher abundance of HSP70 mRNA at each time point after heat stress showed prolonged effect of heat stress in HF PBMCs. The data presented here provided initial evidence of transcriptional differences in PBMCs of different cattle types and buffaloes and warrant further research.

Sublethal heat shock induces premature senescence rather than apoptosis in human mesenchymal stem cells

Stem cells in adult organism are responsible for cell turnover and tissue regeneration. The study of stem cell stress response contributes to our knowledge on the mechanisms of damaged tissue repair. Previously, we demonstrated that sublethal heat shock (HS) induced apoptosis in human embryonic stem cells. This study aimed to investigate HS response of human adult stem cells. Human mesenchymal stem cells (MSCs) cultivated in vitro were challenged with sublethal HS. It was found that sublethal HS did not affect the cell viability assessed by annexin V/propidium staining. However, MSCs subjected to severe HS exhibited features of stress-induced premature senescence (SIPS): irreversible cell cycle arrest, altered morphology, increased expression of senescence-associated β-galactosidase (SA-β-gal) activity, and induction of cyclin-dependent kinase inhibitor p21 protein. High level of Hsp70 accumulation induced by sublethal HS did not return to the basal level, at least, after 72 h of the cell recovery when most cells exhibited SIPS hallmarks. MSCs survived sublethal HS, and resumed proliferation sustained the properties of parental MSCs: diploid karyotype, replicative senescence, expression of the cell surface markers, and capacity for multilineage differentiation. Our results showed for the first time that in human MSCs, sublethal HS induced premature senescence rather than apoptosis or necrosis. MSC progeny that survived sublethal HS manifested stem cell properties of the parental cells: limited replicative life span and multilineage capacity.

Sublethal heat treatment promotes epithelial-mesenchymal transition and enhances the malignant potential of hepatocellular carcinoma

Radiofrequency ablation (RFA) is a potentially curative therapy for hepatocellular carcinoma (HCC). However, incomplete RFA can induce accelerated invasive growth at the periphery. The mechanisms underlying the RFA-induced tumor promotion remain largely unexplored. Three human HCC cell lines were exposed to 45°C-55°C for 10 minutes, simulating the marginal zone of RFA treatment. At 5-12 days post-treatment cell proliferation, parameters of epithelial-mesenchymal transition (EMT), and activation of mitogen-activated protein kinases were analyzed. Livers from patients with viral hepatitis without and with HCC (n = 114) were examined to confirm the relevance of altered kinase patterns. In vivo tumorigenic potential of heat-treated versus untreated HCC cells was studied in nude mice. Heating to 55°C killed all HCC cells, whereas 65%-85% of cells survived 48°C-50°C, developing spindle-like morphology and expressing CD133, cytokeratin (CK)7, CK19, procollagen-α1(I), and Snail at day 5 after heat exposure, which returned to baseline at day 12. Heat-exposed HCC cells showed enhanced proliferation and prominent activation of p46-Shc (Src homology and collagen) and downstream extracellular signal-related kinase (Erk)1/2. In patients, Shc expression correlated with malignant potential and overall survival. Blocking Erk1/2 reduced proliferation and EMT-like changes of heat-treated HCC cells. Implantation of heat-exposed HEPG2 cells into nude mice induced significantly larger, more aggressive tumors than untreated cells. Sublethal heat treatment skews HCC cells toward EMT and transforms them to a progenitor-like, highly proliferative cellular phenotype in vitro and in vivo, which is driven significantly by p46Shc-Erk1/2. Suboptimal RFA accelerates HCC growth and spread by transiently inducing an EMT-like, more aggressive cellular phenotype.

Cytotoxicity and the induction of the stress protein Hsp 70 in Chang liver cells in response to zearalenone-induced oxidative stress

Zearalenone (ZEN) has been implicated in several cases of mycotoxicosis in farm animals and humans. The toxic effects of ZEN have been well characterized, but little is known regarding the mechanisms of ZEN toxicity, including the involvement of the oxidative stress pathway. Using Chang liver cells as a model, the aim of this study was to determine if ZEN could elevate the expression of the heat shock protein Hsp 70, induce cytotoxicity and modulate the levels of glutathione (GSH) and thiobarbituric acid reactive substance (TBARS). In addition, the cytoprotective effects of N-acetylcysteine amide (NACA) pre-treatment were assessed. Finally, the involvement of oxidative stress in ZEN-induced toxicity was confirmed.The results of this study demonstrated that ZEN-induced Hsp 70 expression in a dose- and time-dependent manners. This effect occurred at low-ZEN concentrations, and could therefore be considered a biomarker of ZEN-induced toxicity. The cytotoxicity was reduced when Chang liver cells were exposed to sub-lethal heat shock prior to ZEN treatment, demonstrating a cytoprotective effect of Hsp 70. This cytoprotective effect suggested that Hsp 70 might play a key role in the cellular defense mechanism. When cells were pre-treated with NACA prior to ZEN treatment, the cells were also protected from toxicity. This NACA cytoprotective effect suggested the involvement of oxidative stress in ZEN-induced toxicity, and this mechanism was supported by reduced Hsp 70 expression, inhibited cytolethality, increased GSH levels and decreased TBARS formation when cells were pre-treated with NACA prior to ZEN exposure.Our data clearly demonstrated that ZEN induced cytotoxicity in Chang liver cells by inhibiting cell proliferation, decreasing GSH levels and increasing TBARS formation in a dose-dependent manner. ZEN also, induced Hsp 70 expression, and the side effects of ZEN were significantly alleviated by pre-treatment with NACA. Oxidative stress is likely to be one of the primary pathways of ZEN toxicity. This oxidative stress may contribute, at least in part, to the mechanism of ZEN-induced cytotoxicity.

8:40 AM Abstract No. 190 - Sublethal thermal stress induces accelerated proliferation and growth factor expression in hepatocellular carcinoma

Purpose Tumor recurrence following percutaneous thermal ablation of hepatocellular carcinoma (HCC) remains a significant clinical problem, particularly in tumors ≥3.0cm. The aim of the present study was to test the hypothesis that sublethal heat stress of HCC cells or hepatocytes stimulates accelerated proliferation of non-heat stressed HCC cells and to identify candidate heat-stress induced pro-tumorigenic growth factors. Materials and Methods A small number of HCC cells stably transfected to constitutively express firefly luciferase (N1S1luc and AS30Dluc) were co-cultured with a feeder layer of non-transfected HCC cells (N1S1 or AS30D) or hepatocytes (Clone9) that were heat stressed to induce complete (50 C), partial (45 C) or no (37 C) cell death (N=3). Proliferation of non-heat stressed HCC cells was monitored by non-invasive molecular bioluminescence imaging from 0-6 days using IVIS 200 and a standard curve was generated to estimate cell number in vitro. The two HCC cell lines and the hepatocyte cell line underwent sublethal heat stress (45 C) or control (37 C) and recovered after 6 hours at 37 C for whole transcriptome microarray analysis to assess heat stress induced growth factor expression (N=4). Results Sublethal heat stress (45 C) of both HCC cells and hepatocytes induced a significant increase in proliferation of non-heat stressed N1S1luc (p Conclusion These data suggest that sublethal thermal injury to HCC cells and hepatocytes stimulates proliferation of residual HCC cells and induces a significant increase in numerous pro-tumorigenic growth factors. Thermal ablation induced growth factor signaling warrants further investigation as a mechanism for recurrence and progression following thermal ablation of HCC.

Mechanism of Bacterial Inactivation by (+)-Limonene and Its Potential Use in Food Preservation Combined Processes

This work explores the bactericidal effect of (+)-limonene, the major constituent of citrus fruits' essential oils, against E. coli. The degree of E. coli BJ4 inactivation achieved by (+)-limonene was influenced by the pH of the treatment medium, being more bactericidal at pH 4.0 than at pH 7.0. Deletion of rpoS and exposure to a sub-lethal heat or an acid shock did not modify E. coli BJ4 resistance to (+)-limonene. However, exposure to a sub-lethal cold shock decreased its resistance to (+)-limonene. Although no sub-lethal injury was detected in the cell envelopes after exposure to (+)-limonene by the selective-plating technique, the uptake of propidium iodide by inactivated E. coli BJ4 cells pointed out these structures as important targets in the mechanism of action. Attenuated Total Reflectance Infrared Microspectroscopy (ATR-IRMS) allowed identification of altered E. coli BJ4 structures after (+)-limonene treatments as a function of the treatment pH: β-sheet proteins at pH 4.0 and phosphodiester bonds at pH 7.0. The increased sensitivity to (+)-limonene observed at pH 4.0 in an E. coli MC4100 lptD4213 mutant with an increased outer membrane permeability along with the identification of altered β-sheet proteins by ATR-IRMS indicated the importance of this structure in the mechanism of action of (+)-limonene. The study of mechanism of inactivation by (+)-limonene led to the design of a synergistic combined process with heat for the inactivation of the pathogen E. coli O157:H7 in fruit juices. These results show the potential of (+)-limonene in food preservation, either acting alone or in combination with lethal heat treatments.

Intracranial hyperthermia through local photothermal heating with a fiberoptic microneedle device

The fiberoptic microneedle device (FMD) seeks to leverage advantages of both laser-induced thermal therapy (LITT) and convection-enhanced delivery (CED) to increase volumetric dispersal of locally infused chemotherapeutics through sub-lethal photothermal heat generation. This study focused on determination of photothermal damage thresholds with 1,064 nm light delivered through the FMD into in vivo rat models. FMDs capable of co-delivering laser energy and fluid agents were fabricated through a novel off-center splicing technique involving fusion of a multimode fiberoptic to light-guiding capillary tubing. FMDs were positioned at a depth of 2.5 mm within the cerebrum of male rats with fluoroptic temperature probes placed within 1 mm of the FMD tip. Irradiation (without fluid infusion) was conducted at laser powers of 0 (sham), 100, 200, 500, or 750 mW. Evans blue-serum albumin conjugated complex solution (EBA) and laser energy co-delivery were performed in a second set of preliminary experiments. Maximum, steady-state temperatures of 38.7 ± 1.6 and 42.0 ± 0.9 °C were measured for the 100 and 200 mW experimental groups, respectively. Histological investigation demonstrated needle insertion damage alone for sham and 100 mW irradiations. Photothermal damage was detected at 200 mW, although observable thermal damage was limited to a small penumbra of cerebral cortical microcavitation and necrosis that immediately surrounded the region of FMD insertion. Co-delivery of EBA and laser energy presented increased volumetric dispersal relative to infusion-only controls. Fluoroptic temperature sensing and histopathological assessments demonstrated that a laser power of 100 mW results in sub-lethal brain hyperthermia, and the optimum, sub-lethal target energy range is likely 100-200 mW. The preliminary FMD-CED experiments confirmed the feasibility of augmenting fluid dispersal using slight photothermal heat generation, demonstrating the FMD's potential as a way to increase the efficacy of CED in treating MG.

Evaluating the Predictive Ability of a Path-Dependent Thermal Inactivation Model for Salmonella Subjected to Prior Sublethal Heating in Ground Turkey, Beef, and Pork

Pathogen thermal inactivation models currently available to and used by industry consider only the present state of the product when predicting inactivation rates. However, bacteria subjected to sublethal thermal injury can develop partial protection against lethal temperatures. The objective of this study was to extend the capabilities of a previously published path-dependent Salmonella inactivation model by accounting for longer sublethal heating periods and different substrates and to test this new model against independent data. Ground samples of irradiated (> 10 kGy) turkey breast, beef round, and pork loin were inoculated with an eight-serovar Salmonella cocktail and subjected to 53 nonisothermal treatments (in triplicate) that combined a linear heating rate (1, 2, 3, 4, or 7 K/min), a variable length sublethal holding period (at 40, 45, or 50°C), a lethal holding temperature (55, 58, 61, or 64°C), and a nominal target kill (3- or 5-log reductions) (n = 159 for each meat species). When validated against nonisothermal data from similar treatments, traditional state-dependent model predictions resulted in root mean squared errors (RMSEs) of 2.9, 2.2, and 4.6 log CFU/g for turkey, beef, and pork, respectively. RMSEs for the new path-dependent model were 0.90, 0.81, and 0.82 log CFU/g for the same species, respectively, with reductions in error of 63 to 82 % relative to the state-dependent model. This new path-dependent model can significantly reduce error from the state-dependent model and could become a useful tool for assuring product safety, particularly relative to slow heating processes.