Storage quality of dark, firm, dry meat.

Dark, firm, dry meat contains little or no glucose. Therefore, spoilage bacteria growing on such meat immediately attack amino acids so that spoilage odors and ammonia are detectable at comparatively low cell densities (about 10(6)/cm2).     

Storage quality of dark, firm, dry meat.

Dark, firm, dry meat contains little or no glucose. Therefore, spoilage bacteria growing on such meat immediately attack amino acids so that spoilage odors and ammonia are detectable at comparatively low cell densities (about 10(6)/cm2).     

A note on microbial spoilage of sheep meat at ambient temperature

Shelf-life and microbial spoilage of sheep carcass meat at ambient temperature under commercial conditions were studied. The initial bacterial count of carcasses ranged 5.6-5.8 log/cm2. Staphylococcus spp. (48%) predominated in the initial microflora of carcasses followed by Micrococcus spp. (19%) and Escherichia spp. (12%). Microbial spoilage of carcasses occurred around 20 h when the bacterial count reached 8.0-9.0 log/cm2. Thus the shelf life of carcasses at ambient temperature was 19 h. The predominant micro-organisms at the time of spoilage were Escherichia and 'Acinetobacter-like' organisms. It was also observed that Enterobacter, Pseudomonas and Staphylococcus spp. could form a major part of the final flora. The presence of Escherichia and Staphylococcus spp. in higher percentages on the surface of carcasses at the time of spoilage presents the scope for health hazards.     

The Development of Aerobic Spoilage Flora on Meat Stored at Chill Temperatures

In meat juice medium, aerobic spoilage bacteria utilized the following substrates in the order shown: Pseudomonos , glucose, amino acids, lactic acid; Acinetobacter , amino acids, lactic acid: Enterobacter , glucose, glucose-6-phosphate, amino acids; Microbacterium thermosphactum , glucose, glutamate. All the bacteria grew at their maximum rate utilizing the first and second substrates, but the growth rates declined when these were exhausted. The growth rate of Acinetobacter was reduced at pH 5·7 and below. All other species grew at their maximum rate within the pH range 5·5–7·0. On meat pseudomonads grew faster than the other species at all temperatures between 2° and 15°C. Interactions between any two species were observed only when one organism had attained its maximum cell density. Substrate exhaustion at the meat surface did not limit bacterial growth and it is suggested that the maximum cell density of aerobic spoilage cultures is determined by oxygen limitation of growth.     

Interaction of live and dead Escherichia coli O157:H7 and fluorescent microspheres with lettuce tissue suggests bacterial processes do not mediate adherence

AIMS: The goal of this study was to determine whether any specific bacterial processes (biochemical or genetic) or cell surface moieties were required for the interaction between Escherichia coli O157:H7 and lettuce plant tissue. METHODS AND RESULTS: Escherichia coli O157:H7 and Fluospheres (fluorescent polystyrene microspheres) were used in experiments to investigate interactions with lettuce. Fluospheres were used as they are a non-biological material, of similar size and shape to a bacterial cell, but lack bacterial cell surface moieties and the ability to respond genetically. Live and glutaraldehyde-killed E. coli O157:H7 attached at levels of c. 5.8 log(10) cells per cm(2) following immersion of lettuce pieces into a suspension containing c. 8 log(10) CFU ml(-1). In a separate experiment, numbers of bacteria or Fluospheres associated with lettuce decreased by c. 1.5 log cm(-2) following a 1-min wash. Exposure times of 1 min, 1 h, or 6 h had little effect on the level of attachment for Fluospheres, and live or killed cells of E. coli O157:H7 to lettuce tissue. SIGNIFICANCE: These results indicate that bacterial processes and cell surface moieties are not required for the initial interaction of E. coli O157:H7 to lettuce plant tissue.     

Measurement of bacterial random motility and chemotaxis coefficients: I. Stopped-flow diffusion chamber assay

Bacterial chemotaxis, the directed movement of a cell population in response to a chemical gradient, plays a critical role in the distribution and dynamic interaction of bacterial populations in nonmixed systems. Therefore, in order to make reliable predictions about the migratory behavior of bacteria within the environment, a quantitative characterization of the chemotactic response in terms of intrinsic cell properties is needed.The design of the stopped-flow diffusion chamber (SFDC) provides a well-characterized chemical gradient and reliable method for measuring bacterial migration behavior. During flow through the chamber, a step change in chemical concentration is imposed on a uniform suspension of bacteria. Once flow is stopped, diffusion causes a transient chemical gradient to develop, and bacteria respond by forming a band of high cell density which travels toward higher concentrations of the attractant. Changes in bacterial spatial distributions observed through light scattering are recorded on photomicrographs during a 10-min period. Computer-aided image analysis converts absorbance of the photographic negatives to a digital representation of bacterial density profiles. A mathematical model (part II) is used to quantitatively characterize these observations in terms of intrinsic cell parameters: a chemotactic sensitivity coefficient, mu(0), from the aggregate cell density accumulated in the band and a random motility coefficient, mu, from population dispersion in the absence of a chemical gradient.Using the SFDC assay and an individual-cell-based mathematical model, we successfully determined values for both of these population parameters for Escherichia coli K12 responding to fucose. The values obtained were mu = 1.1 +/- 0. 4 x 10(-5) cm(2)/s and chi(o) = 8 +/- 3 +/- 10(-5) cm(2)/s. We have demonstrated a method capable of determining these parameter values from the now validated mathematical model which will be useful for predicting bacterial migration in application systems.     

Use of a Titrimetric Method to Assess the Bacterial Spoilage of Fresh Beef

A new method of determining bacterial spoilage in fresh beef is presented. The technique is based upon the fact that as beef undergoes refrigerator spoilage, there is a gradual increase in the production of alkaline substances by the spoilage flora. The level of these substances was measured by titrating meat homogenates to a pH 5.00 end point, employing 0.02 n HCl and an autotitrator. When 23 samples of ground beef from retail stores were tested, an average of 1.32 ml of acid was required for titration of 1 g of fresh beef to pH 5.00, whereas 2.58 ml was required for the same meat at the onset of spoilage. Preliminary data indicate that beef which requires more than 2 ml of 0.02 n HCl/g to lower its pH to 5.00 under the conditions of the test is in some state of incipient spoilage. The statistical correlation between titration values, log bacterial numbers, and extract-release volume was high (P < 0.001). The technique is simple to execute and is highly reproducible, and duplicate samples can be run within 15 min.     

Fat liquefaction: effect of low-level laser energy on adipose tissue

Low-level laser energy has been increasingly used in the treatment of a broad range of conditions and has improved wound healing, reduced edema, and relieved pain of various etiologies. This study examined whether 635-nm low-level lasers had an effect on adipose tissue in vivo and the procedural implementation of lipoplasty/liposuction techniques. The experiment investigated the effect of 635-nm, 10-mW diode laser radiation with exclusive energy dispersing optics. Total energy values of 1.2 J/cm(2), 2.4 J/cm(2), and 3.6 J/cm(2) were applied on human adipose tissue taken from lipectomy samples of 12 healthy women. The tissue samples were irradiated for 0, 2, 4, and 6 minutes with and without tumescent solution and were studied using the protocols of transmission electron microscopy and scanning electron microscopy. Nonirradiated tissue samples were taken for reference. More than 180 images were recorded and professionally evaluated. All microscopic results showed that without laser exposure the normal adipose tissue appeared as a grape-shaped node. After 4 minutes of laser exposure, 80 percent of the fat was released from the adipose cells; at 6 minutes of laser exposure, 99 percent of the fat was released from the adipocyte. The released fat was collected in the interstitial space. Transmission electron microscopic images of the adipose tissue taken at x60,000 showed a transitory pore and complete deflation of the adipocytes. The low-level laser energy affected the adipose cell by causing a transitory pore in the cell membrane to open, which permitted the fat content to go from inside to outside the cell. The cells in the interstitial space and the capillaries remained intact. Low-level laser-assisted lipoplasty has a significant impact on the procedural implementation of lipoplasty techniques.     

The proliferative and morphologic responses of a colon carcinoma cell line (LIM 1215) require the production of two autocrine factors.

The role of autocrine growth factors in tumor cell growth has been difficult to prove. Our results indicate that more than one autocrine factor is required for the autonomous growth of the LIM 1215 colonic carcinoma cell line. Furthermore, the morphologic changes induced by epidermal growth factor (EGF) are also density dependent and appear to require a synergistic autocrine factor. The serum-free proliferation of the colonic carcinoma cell line LIM 1215 depends on cell density and the presence of EGF (A. Sizeland, S. Bol, and A.W. Burgess, Growth Factors 4:129-143, 1991). At cell densities below 10(4)/cm2, conditioned medium (from cells at a density of 10(5)/cm2) was required for the cells to elicit a mitogenic response to exogenous EGF. At higher cell densities (10(5)/cm2), the cells were independent of both exogenous EGF and conditioned medium. In addition, the EGF receptor was found to be phosphorylated on tyrosine in LIM 1215 cells proliferating at high density, suggesting that the autocrine production of transforming growth factor alpha (TGF alpha) and subsequent ligation to the EGF receptor was occurring. The proliferation of cells at high density was partly inhibited by TGF alpha antibodies but was almost completely inhibited by an antisense oligonucleotide to TGF alpha. The antisense inhibition could be overcome by the addition of EGF, indicating that the effect of the antisense TGF alpha oligonucleotide was on the production of autocrine TGF alpha. LIM 1215 cells were also observed to undergo morphologic changes (spreading and actin cable organization) in response to EGF. These changes were density dependent, but they occurred with a cell density dependence different from that of the proliferative response. These results suggest two possibilities: that the morphologic changes and proliferative responses have different sensitivities to the autocrine factors or that the actions of the autocrine factors are mediated through different signal transduction pathways.     

Nuclear heterogeneity and proliferative capacity of human adipose derived MSC-like cells

Adipose derived stem cells (ADSCs) are MSC-like cells which could be easily used for regenerative medicine. Here, the morphology and proliferative capacity of human ADSCs is discribed. ADSCs were analyzed after one month of cultivation at a density of 10 cells/cm2. 21 colonies were counted. Few atypical cells (huge nuclei and cytoplasm) were found in 9 out of 17 colonies analyzed. ANOVA demonstrated that colonies also differed (P = 0.0025) in nuclei dimensions and scatter in the dimensions in each colony. Nuclei dimensions and cell density logarithms correlated in reverse proportion (-0.7; P = 0.002). Thus, ADSCs were heterogeneous and represented two types of cells: small highly proliferative and large low proliferative cells. Cell heterogeneity observed in some colonies might be due to cells registered at different cell cycle phases. Stable and typical morphology, colony-formation capability and high proliferative capacity of cells indicate visceral adipose tissue as a rich source of ADSCs.     

Spoilage of chicken skin at 2 degrees C: electron microscopic study.

Microscopic techniques were used in conjunction with normal microbiological procedures to examine the development of the spoilage microflora on the skin of chicken carcasses held at 2 degrees C. Pigmented and nonpigmented psychrotrophic pseudomonads were the major spoilage bacteria isolated at all stages of storage examined. The spoilage microflora grow within a liquid film covering the skin surface, as well as in feather follicle shafts. Penetration and disruption of skin tissue were not observed even after onset of organoleptic spoilage. Bacteria were not attached to the skin by extracellular bridging substances. These data suggest a nonspecific histological-microbiological relationship between he spoilage association and the skin substrate.     

Effect of growth rate and hydrophobicity on bacteria surviving protozoan grazing

Measurements were made of the predation by Tetrahymena thermophila on several bacterial species in media containing heat-killed Escherichia coli cells to serve as an alternative prey. If grazing pressure was initially not intense on a mixture of bacterial species, the species that survived protozoan feeding at greater densities were those that grew quickly before the onset of active predation. If members of several species were incubated individually at similar initial densities with actively grazing T. thermophila, some species survived at ca. 10(4)/ml, some survived at ca. 10(2)/ml, and others were eliminated. Members of the first two groups but not the third group were able to multiply in the medium in the absence of the protozoan, but the growth rates in the protozoan-free medium did not correlate with the number of survivors. However, the species that persisted at the higher densities possessed highly hydrophobic cell surfaces. The size of the surviving population of four bacterial species whose growth was prevented by chloramphenicol correlated with the initial cell density that was incubated with T. thermophila. It is concluded that the individual species surviving predation on a mixture of species is related to the capacity of the bacterium to grow, the hydrophobicity of its cell surface, and the population density of the species before the onset of intense grazing.     

Effect of growth rate and hydrophobicity on bacteria surviving protozoan grazing.

Measurements were made of the predation by Tetrahymena thermophila on several bacterial species in media containing heat-killed Escherichia coli cells to serve as an alternative prey. If grazing pressure was initially not intense on a mixture of bacterial species, the species that survived protozoan feeding at greater densities were those that grew quickly before the onset of active predation. If members of several species were incubated individually at similar initial densities with actively grazing T. thermophila, some species survived at ca. 10(4)/ml, some survived at ca. 10(2)/ml, and others were eliminated. Members of the first two groups but not the third group were able to multiply in the medium in the absence of the protozoan, but the growth rates in the protozoan-free medium did not correlate with the number of survivors. However, the species that persisted at the higher densities possessed highly hydrophobic cell surfaces. The size of the surviving population of four bacterial species whose growth was prevented by chloramphenicol correlated with the initial cell density that was incubated with T. thermophila. It is concluded that the individual species surviving predation on a mixture of species is related to the capacity of the bacterium to grow, the hydrophobicity of its cell surface, and the population density of the species before the onset of intense grazing.     

Bacterial sampling techniques on beef, pork, and lamb adipose tissue.

Two methods were used to evaluate bacterial recovery from beef, pork, and lamb adipose tissue. Higher counts were obtained with a tissue removal and fluid agitation technique (shaking) than with surface swabbing, but only when bacterial levels were low. Bacterial recovery by both methods was unaffected by specie origin of adipose tissue and differences in surface texture, sample storage time (12 versus 6 days), and duration fluid agitation (5 versus 10 min).     

Bacterial sampling techniques on beef, pork, and lamb adipose tissue.

Two methods were used to evaluate bacterial recovery from beef, pork, and lamb adipose tissue. Higher counts were obtained with a tissue removal and fluid agitation technique (shaking) than with surface swabbing, but only when bacterial levels were low. Bacterial recovery by both methods was unaffected by specie origin of adipose tissue and differences in surface texture, sample storage time (12 versus 6 days), and duration fluid agitation (5 versus 10 min).     

Dynamics of microbial communities on marine snow aggregates: colonization,growth, detachment,and grazing mortality of attached bacteria

We studied the dynamics of microbial communities attached to model aggregates (4-mm-diameter agar spheres) and the component processes of colonization, detachment, growth, and grazing mortality. Agar spheres incubated in raw seawater were rapidly colonized by bacteria, followed by flagellates and ciliates. Colonization can be described as a diffusion process, and encounter volume rates were estimated at about 0.01 and 0.1 cm(3) h(-1) for bacteria and flagellates, respectively. After initial colonization, the abundances of flagellates and ciliates remained approximately constant at 10(3) to 10(4) and approximately 10(2) cells sphere(-1), respectively, whereas bacterial populations increased at a declining rate to >10(7) cells sphere(-1). Attached microorganisms initially detached at high specific rates of approximately 10(-2) min(-1), but the bacteria gradually became irreversibly attached to the spheres. Bacterial growth (0 to 2 day(-1)) was density dependent and declined hyperbolically when cell density exceeded a threshold. Bacterivorous flagellates grazed on the sphere surface at an average saturated rate of 15 bacteria flagellate(-1) h(-1). At low bacterial densities, the flagellate surface clearance rate was approximately 5 x 10(-7) cm(2) min(-1), but it declined hyperbolically with increasing bacterial density. Using the experimentally estimated process rates and integrating the component processes in a simple model reproduces the main features of the observed microbial population dynamics. Differences between observed and predicted population dynamics suggest, however, that other factors, e.g., antagonistic interactions between bacteria, are of importance in shaping marine snow microbial communities.     

Optimization of DNA immobilization on gold electrodes for label-free detection by electrochemical impedance spectroscopy

The ability to immobilize DNA probes onto gold substrates at an optimum surface density is key in the development of a wide range of DNA biosensors. We present a method to accurately control probe DNA surface density by the simultaneous co-immobilization of thiol modified probes and mercaptohexanol. Probe surface density is controlled by the thiol molar ratio in solution, with a linear relationship between thiol molar ratio and probe density spanning (1-9) x10(12)/cm2. The probe surface density per microscopic surface area was determined using chronocoulometry, and a detailed analysis of the method presented. Using this sample preparation method, the effect of probe density and hybridization on the charge transfer resistance with the negatively charged ferri/ferrocyanide redox couple was determined. Above a threshold probe surface density of 2.5 x 10(12)/cm2, electrostatic repulsion from the negatively charged DNA modulates the charge transfer resistance, allowing hybridization to be detected. Below the threshold density no change in charge transfer resistance with probe density or with hybridization occurs. The probe surface density was optimized to obtain the maximum percentage change in charge transfer resistance with hybridization.     

Stromelysin-2 (MMP-10) deficiency does not affect adipose tissue formation in a mouse model of nutritionally induced obesity

Adipose tissue development is associated with angiogenesis, adipogenesis and extracellular matrix degradation. The class of matrix metalloproteinases contributes to these processes, but little information is available on the role of individual proteinases. We report that stromelysin-2 (MMP-10) deficiency has no significant effect on total body weight or on subcutaneous (SC) or gonadal (GON) adipose tissue mass of mice kept on a high fat diet for 15 weeks. The adipocyte size and density in SC and GON adipose tissues were also comparable in MMP-10 deficient and wild-type control mice. Similarly, blood vessel size and density in obese SC and GON adipose tissues was not affected by MMP-10 deficiency.Metabolic parameters and blood cell composition were similar for both genotypes. Stromelysin-1 (MMP-3) expression was significantly reduced in adipose tissues of the deficient mice as compared to the wild-type controls.These data indicate that MMP-10 does not significantly contribute to adipose tissue development and associated angiogenesis in a mouse model of nutritionally induced obesity.     

Presence of functional TLR2 and TLR4 on human adipocytes

In addition to the well-known role of adipose tissue in energy metabolism, it has recently been demonstrated that this tissue can secrete a large array of molecules, including inflammatory cytokines. Furthermore, recent studies suggest that adipose cells can behave as immune cells. Therefore, the aim of this study was to determine the presence of the two most prominent ‘pattern recognition receptors’ for bacterial and fungal cell wall components, TLR2 and TLR4 on human adipose cells, as well as to assess their functionality. We demonstrated that TLR2 and TLR4 were expressed at relatively high levels (compared to a monocyte cell line) on the surface of human adipose cells. Stimulation of human adipocytes with lipopolysaccharide (LPS), or with lipoteichoic acid (LTA), two specific ligands of TLR4 and TLR2, respectively, induced a strong increase in TNFα production. The specificity of the response was demonstrated by the use of anti-TLR4 and anti-TLR2 blocking antibodies, which were able to decrease LPS- or LTA-induced TNFα secretion. Thus, it is clear that these receptors are functional in human adipocytes. This study adds weight to the argument that human fat tissue plays a potential role in innate immunity. Sandrine Bés-Houtmann, Régis Roche, Christian Lefebvre d’Hellencourt and Maya Cesari have contributed equally to this work.