Comparison of Natural Killer Cells Differentiated from Various Pluripotent Stem Cells.

Allogeneic natural killer (NK) cell therapy has been effective in treating cancer. Many studies have tested NK cell therapy using human pluripotent stem cells (hPSCs). However, the impacts of the origin of PSC-NK cells on competence are unclear. In this study, several types of hPSCs, including human-induced PSCs (hiPSCs) generated from CD34+, CD3-CD56+, and CD56- cells in umbilical cord blood (UCB), three lines of human embryonic stem cells (hESCs, ES-1. ES-2 and ES-3) and MHC I knockout (B2M-KO)-ESCs were used to differentiate into NK cells and their capacities were analyzed. All PSC types could differentiate into NK cells. Among the iPSC-derived NK cells (iPSC-NKs) and ESC-derived NK cells (ES-NKs), 34+ iPSCs and ES-3 had a higher growth rate and cytotoxicity, respectively, ES-3 also showed better efficacy than 34+ iPSCs. B2M-KO was similar to the wild type. These results suggest that the screening for differentiation of PSCs into NK cells prior to selecting the PSC lines for the development of NK cell immunotherapy is an essential process for universal allotransplantation, including the chimeric antigen receptor (CAR).

Intrinsic Magnetic Order of Chemically Exfoliated 2D Ruddlesden-Popper Organic-Inorganic Halide Perovskite Ultrathin Films.

Thin film fabrication of 2D layered organic-inorganic hybrid perovskites (2D-OIHPs) for spintronic applications has been attempted using solution-based process like Langmuir-Blodgett technique. However, monolayer or few-layered 2D magnets are not yet realized, even though a wide spectrum of 2D Ruddlesden-Popper (RP) OIHPs are known as quasi-2D Heisenberg magnets in bulk compounds. Here, chemical exfoliation by solvent engineering is applied to successfully synthesize large-sized, few unit-cell-thick 2D RP-OIHPs. Comprehensive structural characterization reveals that binary co-solvents with high relative polarity in spin coating technique are the most effective among nine kinds of solvents. Above all, this enables few-layered 2D RP-OIHP ultrathin films sustaining their intrinsic magnetic order. It is found that XY-like magnetic anisotropy driven by Jahn-Teller effect responsible for ferromagnetism in seven-layered (C6 H5 CH2 CH2 NH3 )2 CuCl4 ultrathin films remains very robust, whereas Ising-like dipolar anisotropy responsible for canted antiferromagnetism in ten-layered (C6 H5 CH2 CH2 NH3 )2 MnCl4 ultrathin films is greatly reduced. It is expected that ferromagnetism even at monolayer limit should be possible by means of further sophisticated solvent engineering as long as Jahn-Teller effect is active. The chemical exfoliation using solvent engineering unambiguously can bring about a new breakthrough in the development of 2D RP-OIHP van der Waals magnets for ultrahigh energy-efficient spintronic, opto-spintronic devices.

Butyric acid production from red algae by a newly isolated Clostridium sp. S1.

OBJECTIVE: To produce butyric acid from red algae such as Gelidium amansii in which galactose is a main carbohydrate, microorganisms utilizing galactose and tolerating inhibitors in hydrolysis including levulinic acid and 5-hydroxymethylfurfural (HMF) are required. RESULTS: A newly isolated bacterium, Clostridium sp. S1 produced butyric acid not only from galactose as the sole carbon source but also from a mixture of galactose and glucose through simultaneous utilization. Notably, Clostridium sp. S1 produced butyric acid and a small amount of acetic acid with the butyrate:acetate ratio of 45.4:1 and it even converted acetate to butyric acid. Clostridium sp. S1 tolerated 0.5-2 g levulinic acid/l and recovered from HMF inhibition at 0.6-2.5 g/l, resulting in 85-92% butyric acid concentration of the control culture. When acid-pretreated G. amansii hydrolysate was used, Clostridium sp. S1 produced 4.83 g butyric acid/l from 10 g galactose/l and 1 g glucose/l. CONCLUSION: Clostridium sp. S1 produces butyric acid from red algae due to its characteristics in sugar utilization and tolerance to inhibitors, demonstrating its advantage as a red algae-utilizing microorganism.

Morphology control of surfactant-assisted graphene oxide films at the liquid-gas interface.

Control of a two-dimensional (2D) structure of assembled graphene oxide (GO) sheets is highly desirable for fundamental research and potential applications of graphene devices. We show that an alkylamine surfactant, i.e., octadecylamine (ODA), Langmuir monolayer can be utilized as a template for adsorbing highly hydrophilic GO sheets in an aqueous subphase at the liquid-gas interface. The densely packed 2-D monolayer of such complex films was obtained on arbitrary substrates by applying Langmuir-Schaefer or Langmuir-Blodgett technique. Morphology control of GO sheets was also achieved upon compression by tuning the amount of spread ODA molecules. We found that ODA surfactant monolayers prevent GO sheets from sliding, resulting in formation of wrinkling rather than overlapping at the liquid-gas interface during the compression. The morphology structures did not change after a graphitization procedure of chemical hydrazine reduction and thermal annealing treatments. Since morphologies of graphene films are closely correlated to the performance of graphene-based materials, the technique employed in this study can provide a route for applications requiring wrinkled graphenes, ranging from nanoelectronic devices to energy storage materials, such as supercapacitors and fuel cell electrodes.

Monophosphoryl lipid A and poly I:C combination enhances immune responses of equine influenza virus vaccine.

Equine influenza is a contagious respiratory disease caused by H3N8 type A influenza virus. Vaccination against equine influenza is conducted regularly; however, infection still occurs globally because of the short immunity duration and suboptimal efficacy of current vaccines. Hence the objective of this study was to investigate whether an adjuvant combination can improve immune responses to equine influenza virus (EIV) vaccines. Seventy-two mice were immunized with an EIV vaccine only or with monophosphoryl lipid A (MPL), polyinosinic-polycytidylic acid (Poly I:C), or MPL + Poly I:C. Prime immunization was followed by boost immunization after 2 weeks. Mice were euthanized at 4, 8, and 32 weeks post-prime immunization, respectively. Sera were collected to determine humoral response. Bone marrow, spleen, and lung samples were harvested to determine memory cell responses, antigen-specific T-cell proliferation, and lung viral titers. MPL + Poly I:C resulted in the highest IgG, IgG1, and IgG2a antibodies and hemagglutination inhibition titers among the groups and sustained their levels until 32 weeks post-prime immunization. The combination enhanced memory B cell responses in the bone marrow and spleen. At 8 weeks post-prime immunization, the combination induced higher CD8+ central memory T cell frequencies in the lungs and CD8+ central memory T cells in the spleen. In addition, the combination group exhibited enhanced antigen-specific T cell proliferation, except for CD4+ T cells in the lungs. Our results demonstrated improved immune responses when using MPL + Poly I:C in EIV vaccines by inducing enhanced humoral responses, memory cell responses, and antigen-specific T cell proliferation.

Targeted sampling reduces the uncertainty in force of infection estimates from serological surveillance.

Age bins are frequently used in serological studies of infectious diseases in wildlife to deal with uncertainty in the age of sampled animals. This study analyzed how age binning and targeted sampling in serological surveillance affect the width of the 95% confidence interval (CI) of the estimated force of infection (FOI) of infectious diseases. We indicate that the optimal target population with the narrowest 95% CI differs depending on the expected FOI using computer simulations and mathematical models. In addition, our findings show that we can substantially reduce the number of animals required to infer transmission risk by tailoring targeted, age-based sampling to specific epidemiological situations.

Clustering of genes from microarray data using hierarchical projective adaptive resonance theory: a case study of tuberculosis.

We propose the hierarchical Projective Adaptive Resonance Theory (PART) algorithm for classification of gene expression data. This algorithm is realized by combing transposed quasi-supervised PART and unsupervised PART. We develop the corresponding validation statistics for each process and compare it with other clustering algorithms in a case study of tuberculosis (TB). First, we use sample-based transposed quasi-supervised PART to obtain optimal clustering results of samples distinguished by time post-infection and the representative genes for each cluster including up-regulated, down-regulated and stable genes. The up- and down-regulated genes show more than 90% similarity to the result derived from Linear Models for Microarray Data and are verified by weighted k-nearest neighbor model on TB projection. Second, we use gene-based unsupervised PART algorithm to cluster these representative genes where functional enrichment analysis is conducted in each cluster. We further confirm the main immune response of human macrophage-like THP-1 cells against TB within 2 days is type I interferon-mediated innate immunity. This study demonstrates how hierarchical PART algorithm analyzes microarray data. The sample-based quasi-supervised PART extracts representative genes and narrows down the shortlist of disease-relevant genes and gene-based unsupervised PART classifies representative genes that help to interpret immune response against TB.

Association of bacterial isolates and antimicrobial susceptibility between prostatic fluid and urine samples in canine prostatitis with concurrent cystitis.

Most old, intact male dogs usually have prostate disorders, especially benign prostatic hypertrophy and prostatitis with or without abscesses, and concurrent cystitis. The successful treatment of dogs with prostatitis concurrent with cystitis has relied on choosing an appropriate antimicrobial drug based on a bacterial culture and drug sensitivity testing. The objective of the study was to compare the prevalence of bacterial species and results of drug susceptibility testing of bacteria that were isolated from the prostatic fluids and urine samples that were collected from dogs with both prostatitis and cystitis. One hundred and sixty intact male dogs, who presented with both diseases, were recruited for the study. The disease diagnoses were based on clinical history notes, physical examinations, abdominal ultrasonography, prostatic fluid cytology, urinalysis and bacterial cultures from both prostatic fluid and urine samples. The bacterial culture results demonstrated that the major species that were detected in either the prostatic fluid or urine samples were Staphylococcus spp., Escherichia coli, Pseudomonas spp., Streptococcus spp., Proteus mirabilis and Klebsiella pneumoniae. Staphylococcus spp. (26.5 %, 43/162) and Escherichia coli (26.1 %, 12/46) were the most prevalent species from the prostatic fluid and urine samples, respectively. Statistical tests revealed that there were no significantly different prevalence levels among the isolated bacteria between the prostatic fluid and urine samples. Imipenem and gentamicin were the most potent antimicrobial drugs tested against the bacterial isolates in the present study. However, the administration of imipenem to treat prostatitis and cystitis in dogs was of concern. Interestingly, there were no significant differences in the antimicrobial drug susceptibility trends between the prostatic fluid and urine samples. Based on these results, a urine sample might be considered as an optional sample for bacterial cultures and antimicrobial drug susceptibility testing when it is not possible to collect a prostatic fluid sample.

Enhanced butyric acid production using mixed biomass of brown algae and rice straw by Clostridium tyrobutyricum ATCC25755.

A brown alga Saccharina japonica and rice straw are attractive feedstock for microbial butyric acid production. However, inefficient fermentation of mannitol (a dominant component in S. japonica) and toxicity of inhibitors in lignocellulosic hydrolysate are limitations. This study demonstrated that mixed biomass with S. japonica and rice straw was effective in butyric acid production over those restrictions. Mannitol was consumed only when acetic acid was present. Notably, acetic acid was not produced but consumed along with mannitol. By mixing S. japonica and rice straw to take advantage of glucose and acetic acid in rice straw, Clostridium tyrobutyricum effectively consumed mannitol by utilizing acetic acid in hydrolysate and acetic acid derived from glucose with the enhanced butyric acid production. Furthermore, cell growth was restored owing to the decreased inhibitor concentration. The results demonstrate the potential of butyric acid production from mixed biomass of macroalgae/lignocellulose overcoming the drawbacks of single biomass.

Purification and characterization of thermostable beta-glucosidase from the brown-rot basidiomycete Fomitopsis palustris grown on microcrystalline cellulose.

An extracellular beta-glucosidase was purified 154-fold to electrophoretic homogeneity from the brown-rot basidiomycete Fomitopsis palustris grown on 2.0% microcrystalline cellulose. SDS-polyacrylamide gel electrophoresis gel gave a single protein band and the molecular mass of purified enzyme was estimated to be approximately 138 kDa. The amino acid sequences of the proteolytic fragments determined by nano-LC-MS/MS suggested that the protein has high homology with fungal beta-glucosidases that belong to glycosyl hydrolase family 3. The Kms for p-nitorophenyl-beta-D-glucoside (p-NPG) and cellobiose hydrolyses were 0.117 and 4.81 mM, and the Kcat values were 721 and 101.8 per sec, respectively. The enzyme was competitively inhibited by both glucose (Ki= 0.35 mM) and gluconolactone (Ki= 0.008 mM), when p-NPG was used as substrate. The optimal activity of the purified beta-glucosidase was observed at pH 4.5 and 70 degrees. The F. palustris protein exhibited half-lives of 97 h at 55 degrees and 15 h at 65 degrees, indicating some degree of thermostability. The enzyme has high activity against p-NPG and cellobiose but has very little or no activity against p-nitrophenyl-beta-lactoside, p-nitrophenyl-beta-xyloside, p-nitrophenyl-alpha-arabinofuranoside, xylan, and carboxymethyl cellulose. Thus, our results revealed that the beta-glucosidase from F. palustris can be classified as an aryl-beta-glucosidase with cellobiase activity.

Functional analysis of a gene encoding endoglucanase that belongs to glycosyl hydrolase family 12 from the brown-rot basidiomycete Fomitopsis palustris.

The brown-rot basidiomycete Fomitopsis palustris is known to degrade crystalline cellulose (Avicel) and produce three major cellulases, exoglucanases, endoglucanases, and beta- glucosidases. A gene encoding endoglucanase, designated as cel12, was cloned from total RNA prepared from F. palustris grown at the expense of Avicel. The gene encoding Cel12 has an open reading frame of 732 bp, encoding a putative protein of 244 amino acid residues with a putative signal peptide residing at the first 18 amino acid residues of the N-terminus of the protein. Sequence analysis of Cel12 identified three consensus regions, which are highly conserved among fungal cellulases belonging to GH family 12. However, a cellulose-binding domain was not found in Cel12, like other GH family 12 fungal cellulases. Northern blot analysis showed a dramatic increase of cel12 mRNA levels in F. palustris cells cultivated on Avicel from the early to late stages of growth and the maintenance of a high level of expression in the late stage, suggesting that Cel12 takes a significant part in endoglucanase activity throughout the growth of F. palustris. Adventitious expression of cel12 in the yeast Pichia pastoris successfully produced the recombinant protein that exhibited endoglucanase activity with carboxymethyl cellulose, but not with crystalline cellulose, suggesting that the enzyme is not a processive endoglucanase unlike two other endoglucanases previously identified in F. palustris.

Aerobic and anaerobic cellulose utilization by Paenibacillus sp. CAA11 and enhancement of its cellulolytic ability by expressing a heterologous endoglucanase.

For cost-effective lignocellulosic biofuel/chemical production, consolidated bioprocessing (CBP)-enabling microorganisms utilizing cellulose as well as producing biofuel/chemical are required. A novel strain Paenibacillus sp. CAA11 isolated from sediment was found to be not only as a cellulose degrader under both aerobic and strict anaerobic conditions but also as a producer of cellulosic biofuel/chemicals. Paenibacillus sp. CAA11 secreted cellulolytic enzymes by its own secretion system and produced ethanol as well as short-chain organic acids (formic acid, acetic acid, lactic acid) from cellulose. Cellulolytic activity of the strain was significantly enhanced by expressing a heterologous endoglucanase 168Cel5 from Bacillus subtilis under both aerobic and anaerobic conditions. The strain harboring the 168cel5 gene revealed 2-fold bigger halo zone on Congo-red plate and 1.75-fold more aerobic cellulose utilization in liquid medium compared with the negative control. Notably, under anaerobic conditions, the recombinant strain expressing 168Cel5 consumed 1.83-fold more cellulose (5.10 g/L) and produced 5-fold more ethanol (0.65 g/L) along with 5-fold more total acids (1.6 g/L) compared with the control, resulting 2.73-fold higher yields. This result demonstrates the potential of Paenibacillus sp. CAA11 as a suitable aerobic and anaerobic CBP-enabling microbe with cellulolytic production of ethanol and short-chain organic acids.

Solvent-dependent self-assembly of two dimensional layered perovskite (C6H5CH2CH2NH3)2MCl4 (M = Cu, Mn) thin films in ambient humidity.

Two dimensional layered organic-inorganic halide perovskites offer a wide variety of novel functionality such as solar cell and optoelectronics and magnetism. Self-assembly of these materials using solution process (ex. spin coating) makes crystalline thin films synthesized at ambient environment. However, flexibility of organic layer also poses a structure stability issue in perovskite thin films against environment factors (ex. moisture). In this study, we investigate the effect of solvents and moisture on structure and property in the (C6H5(CH2)2NH3)2(Cu, Mn)Cl4 (Cu-PEA, Mn-PEA) perovskite thin films spin-coated on Si wafer using three solvents (H2O, MeOH, MeOH + H2O). A combination of x-ray diffraction (XRD) and x-ray absorption spectroscopy (XAS) show that relative humidity (RH) has a profound effect on perovskite thin films during sample synthesis and storage, depending on the kind of solvent used. The ones prepared using water (Cu-PEA:H2O, Mn-PEA:H2O) show quite different behavior from the other cases. According to time-dependent XRD, reversible crystalline-amorphous transition takes place depending on RH in the former cases, whereas the latter cases relatively remain stable. It also turns out from XAS that Mn-PEA thin films prepared with solvents such as MeOH and MeOH + H2O are disordered to the depth of about 4 nm from surface.

Inferring epidemiological dynamics of infectious diseases using Tajima's D statistic on nucleotide sequences of pathogens.

The estimation of the basic reproduction number is essential to understand epidemic dynamics, and time series data of infected individuals are usually used for the estimation. However, such data are not always available. Methods to estimate the basic reproduction number using genealogy constructed from nucleotide sequences of pathogens have been proposed so far. Here, we propose a new method to estimate epidemiological parameters of outbreaks using the time series change of Tajima's D statistic on the nucleotide sequences of pathogens. To relate the time evolution of Tajima's D to the number of infected individuals, we constructed a parsimonious mathematical model describing both the transmission process of pathogens among hosts and the evolutionary process of the pathogens. As a case study we applied this method to the field data of nucleotide sequences of pandemic influenza A (H1N1) 2009 viruses collected in Argentina. The Tajima's D-based method estimated basic reproduction number to be 1.55 with 95% highest posterior density (HPD) between 1.31 and 2.05, and the date of epidemic peak to be 10th July with 95% HPD between 22nd June and 9th August. The estimated basic reproduction number was consistent with estimation by birth-death skyline plot and estimation using the time series of the number of infected individuals. These results suggested that Tajima's D statistic on nucleotide sequences of pathogens could be useful to estimate epidemiological parameters of outbreaks.

Host-Specific and Segment-Specific Evolutionary Dynamics of Avian and Human Influenza A Viruses: A Systematic Review.

Understanding the evolutionary dynamics of influenza viruses is essential to control both avian and human influenza. Here, we analyze host-specific and segment-specific Tajima's D trends of influenza A virus through a systematic review using viral sequences registered in the National Center for Biotechnology Information. To avoid bias from viral population subdivision, viral sequences were stratified according to their sampling locations and sampling years. As a result, we obtained a total of 580 datasets each of which consists of nucleotide sequences of influenza A viruses isolated from a single population of hosts at a single sampling site within a single year. By analyzing nucleotide sequences in the datasets, we found that Tajima's D values of viral sequences were different depending on hosts and gene segments. Tajima's D values of viruses isolated from chicken and human samples showed negative, suggesting purifying selection or a rapid population growth of the viruses. The negative Tajima's D values in rapidly growing viral population were also observed in computer simulations. Tajima's D values of PB2, PB1, PA, NP, and M genes of the viruses circulating in wild mallards were close to zero, suggesting that these genes have undergone neutral selection in constant-sized population. On the other hand, Tajima's D values of HA and NA genes of these viruses were positive, indicating HA and NA have undergone balancing selection in wild mallards. Taken together, these results indicated the existence of unknown factors that maintain viral subtypes in wild mallards.

Effective isopropanol-butanol (IB) fermentation with high butanol content using a newly isolated Clostridium sp. A1424.

BACKGROUND: Acetone-butanol-ethanol fermentation has been studied for butanol production. Alternatively, to achieve acetone-free butanol production, use of clostridium strains producing butanol and 1,3-propanediol (1,3-PDO) from glycerol, natural and engineered isopropanol-butanol-ethanol (IBE) producers has been attempted; however, residual 1,3-PDO and acetone, low IBE production by natural IBE producers, and complicated gene modification are limitations. RESULTS: Here, we report an effective isopropanol and butanol (IB) fermentation using a newly isolated Clostridium sp. A1424 capable of producing IB from various substrates with a small residual acetone. Notably, this strain also utilized glycerol and produced butanol and 1,3-PDO. After 46.35 g/L of glucose consumption at pH 5.5-controlled batch fermentation, Clostridium sp. A1424 produced 9.43 g/L of butanol and 13.92 g/L of IB at the productivity of 0.29 and 0.44 g/L/h, respectively, which are the highest values in glucose-based batch fermentations using natural IB producers. More interestingly, using glucose-glycerol mixtures at ratios ranging from 20:2 to 14:8 led to not only acetone-free and 1,3-PDO-free IB fermentation but also enhanced IB production along with a much higher butanol content (butanol/isopropanol ratio of 1.81 with glucose vs. 2.07-6.14 with glucose-glycerol mixture). Furthermore, when the mixture of glucose and crude glycerol at the ratio of 14:8 (total concentration of 35.68 g/L) was used, high butanol/isopropanol ratio (3.44) and butanol titer (9.86 g/L) were achieved with 1.4-fold enhanced butanol yield (0.28 g/g) and productivity (0.41 g/L/h) compared to those with glucose only at pH 5.5. CONCLUSIONS: A newly isolated Clostridium sp. A1424 was able to produce butanol and isopropanol from various carbon sources. The productivity and titer of butanol and total alcohol obtained in this study were higher than the previously reported results obtained using other natural IB producers. Use of the mixture of glucose and glycerol was successful to achieve acetone-free, 1,3-PDO-free, and enhanced IB production with higher yield, productivity, and selectivity of butanol compared to those with glucose only, providing great advantages from the perspective of carbon recovery to alcohols. This notable result could be accomplished by isolating an effective IB producer Clostridium sp. A1424 as well as by utilizing glucose-glycerol mixtures.

Butyric acid production from softwood hydrolysate by acetate-consuming Clostridium sp. S1 with high butyric acid yield and selectivity.

The aim of this work was to study the butyric acid production from softwood hydrolysate by acetate-consuming Clostridium sp. S1. Results showed that Clostridium sp. S1 produced butyric acid by simultaneously utilizing glucose and mannose in softwood hydrolysate and, more remarkably, it consumed acetic acid in hydrolysate. Clostridium sp. S1 utilized each of glucose, mannose, and xylose as well as mixed sugars simultaneously with partially repressed xylose utilization. When softwood (Japanese larch) hydrolysate containing glucose and mannose as the main sugars was used, Clostridium sp. S1 produced 21.17g/L butyric acid with the yield of 0.47g/g sugar and the selectivity of 1 (g butyric acid/g total acids) owing to the consumption of acetic acid in hydrolysate. The results demonstrate potential of Clostridium sp. S1 to produce butyric acid selectively and effectively from hydrolysate not only by utilizing mixed sugars simultaneously but also by converting acetic acid to butyric acid.

Electrochemical detoxification of phenolic compounds in lignocellulosic hydrolysate for Clostridium fermentation.

Lignocellulosic biomass is being preferred as a feedstock in the biorefinery, but lignocellulosic hydrolysate usually contains inhibitors against microbial fermentation. Among these inhibitors, phenolics are highly toxic to butyric acid-producing and butanol-producing Clostridium even at a low concentration. Herein, we developed an electrochemical polymerization method to detoxify phenolic compounds in lignocellulosic hydrolysate for efficient Clostridium fermentation. After the electrochemical detoxification for 10h, 78%, 77%, 82%, and 94% of p-coumaric acid, ferulic acid, vanillin, and syringaldehyde were removed, respectively. Furthermore, 71% of total phenolics in rice straw hydrolysate were removed without any sugar-loss. Whereas the cell growth and metabolite production of Clostridium tyrobutyricum and Clostridium beijerinckii were completely inhibited in un-detoxified hydrolysate, those in detoxifying rice straw hydrolysate were recovered to 70-100% of the control cultures. The electrochemical detoxification method described herein provides an efficient strategy for producing butanol and butyric acid through Clostridium fermentation with lignocellulosic hydrolysate.

Antibiotic Resistance and Virulence Potentials of Shiga Toxin-Producing Escherichia coli Isolates from Raw Meats of Slaughterhouses and Retail Markets in Korea.

In this study, the prevalence of Shiga toxin-producing Escherichia coli (STEC) was investigated among raw meat or meat products from slaughterhouses and retail markets in South Korea, and their potential for antibiotic resistance and virulence was further analyzed. A total of 912 raw meats, including beef, pork, and chicken, were collected from 2008 to 2009. E. coli strains were frequently isolated in chicken meats (176/233, 75.9%), beef (102/217, 42.3%), and pork (109/235, 39.2%). Putative STEC isolates were further categorized, based on the presence or absence of the Shiga toxin (stx) genes, followed by standard O-serotyping. Polymerase chain reaction assays were used to detect the previously defined virulence genes in STEC, including Shiga toxins 1 and Shiga toxin 2 (stx1 and 2), enterohemolysin (ehxA), intimin (eaeA), STEC autoagglutination adhesion (saa), and subtilase cytotoxin (subAB). All carried both stx1 and eae genes, but none of them had the stx2, saa, or subAB genes. Six (50.0%) STEC isolates possessed the ehxA gene, which is known to be encoded by the 60-megadalton virulence plasmid. Our antibiogram profiling demonstrated that some STEC strains, particularly pork and chicken isolates, displayed a multiple drug-resistance phenotype. RPLA analysis revealed that all the stx1-positive STEC isolates produced Stx1 only at the undetectable level. Altogether, these results imply that the locus of enterocyte and effacement (LEE)-positive strains STEC are predominant among raw meats or meat products from slaughterhouses or retail markets in Korea.

Effect of temperature on bacterial emissions in composting of swine manure.

Swine manure was subjected to laboratory scale composting in order to quantify bioaerosols, i.e., airborne culturable bacteria and endotoxin, in the exhaust gas, which provided details on the effect of temperature on bacterial emissions. The concentration of airborne bacteria reached 31,250 colony-forming units (CFU)/m(3) during the thermophilic stage of composting, and positively correlated with the temperature profile of the compost pile. Initially, the endotoxin concentration was 1820 endotoxin units (EU)/m(3), but it decreased exponentially as the composting process proceeded. The temperature can be an excellent indicator of bacterial emissions during the composting process, indicating that the composting process requires a consistently high temperature to ensure sanitization of both compost and bacterial emissions. The cumulative emission data showed that emission factors was 11.2-13.5 CFU/g dry swine manure and that of endotoxin was 0.5-0.9 EU/g dry swine manure. The bacterial diversity in the bioaerosol was analyzed by polymerase chain reaction-denaturing gradient gel electrophoresis, revealing the presence of various gram-negative bacterial consortia.