In Vitro and In Silico Characterization of N-Formylated Two-Peptide Bacteriocin from Enterococcus faecalis CAUM157 with Anti-Listeria Activity.

Enterococcus faecalis CAUM157 (KACC 81148BP), a Gram-positive bacteria isolated from raw cow's milk, was studied for its bacteriocin production. The antimicrobial activity of CAUM157 was attributed to a two-peptide class IIb bacteriocin with potent activity against food-borne pathogen Listeria monocytogenes and periodontal disease-causing pathogens (Prevotella intermedia KCTC 15693 T and Fusobacterium nucleatum KCTC 2488 T). M157 bacteriocins exhibit high temperature and pH stability and resist hydrolytic enzyme degradation and detergent denaturation, potentially due to their structural conformation. Based on amino acid sequence, M157A and M157B were predicted to be 5.176 kDa and 5.182 kDa in size, respectively. However, purified bacteriocins and chemically synthesized N-formylated M157 peptides both showed 5.204 kDa (M157A) and 5.209 kDa (M157B) molecular mass, confirming the formylation of the N-terminal methionine of both peptides produced by strain CAUM157. Furthermore, the strain demonstrated favorable growth and fermentation with minimal bacteriocin production when cultured in whey-based media, whereas a 1.0% tryptone or soytone supplementation resulted in higher bacteriocin production. Although Ent. faecalis CAUM157 innately harbors genes for virulence factors and antimicrobial resistance (e.g., tetracycline and erythromycin), its bacteriocin production is valuable in circumventing the need for live microorganisms, particularly in food applications for pathogen control.

Complete genome sequence of bacteriocin-producing Ligilactobacillus salivarius B4311 isolated from fecal samples of broiler chicken with anti-listeria activity.

Ligilactobacillus is a genus of Gram-positive lactobacilli commonly found in the intestinal tracts of vertebrates. It has been granted a Qualified Presumption of Safety (QPS) status from the European Food Safety Authority (EFSA). One specific strain, Ligilactobacillus salivarius B4311, was isolated from fecal samples of broiler chickens from a farm associated with Chung-Ang University (Anseong, Korea). This strain was observed to have inhibitory effects against Listeria monocytogenes. In this paper, we present the complete genome sequence of Lig. salivarius B4311. The whole genome of strain B4311 comprises 2,071,255 bp assembled into 3 contigs representing a chromosome, repA-type megaplasmid, and small plasmid. The genome contains 1,963 protein-coding sequences, 22 rRNA genes, and 78 tRNA genes, with a guanine + cytosine (GC) content of 33.1%. The megaplasmid of strain B4311 was found to contain the bacteriocin gene cluster for salivaricin P, a two-peptide bacteriocin belonging to class IIb.

Detection and characterization of potential virulence determinants in Staphylococcus pseudintermedius and S. schleiferi strains isolated from canine otitis externa in Korea.

BACKGROUND: A recent increase in the occurrence of canine skin and soft tissue infections, including otitis externa and pyoderma, caused by antimicrobial-resistant Staphylococcus pseudintermedius and S. schleiferi has become a significant public and veterinary health issues. OBJECTIVE: We investigated the virulence potentials associated with the occurrence of canine otitis externa in S. pseudintermedius and S. schleiferi. METHODS: In this study, the prevalence of genes encoding leukocidins, exfoliative toxins, and staphylococcal enterotoxins (SEs) was investigated using previously characterized S. pseudintermedius (n = 26) and S. schleiferi (n = 19) isolates derived from canine otitis externa. Susceptibility to cathelicidins (K9CATH and PMAP-36) and hydrogen peroxide (H2O2) was also examined in both staphylococcal species. RESULTS: A high prevalence of genes encoding leukocidins (lukS/F-I, lukS1/F1-S, and lukS2/F2-S), exfoliative toxins (siet, expB, and sset), and SEs was identified in both S. pseudintermedius and S. schleiferi isolates. Notably, S. pseudintermedius isolates possessed higher number of SE genes, especially newer SE genes, than S. schleiferi isolates harboring egc clusters. Although no significant differences in susceptibility to K9CATH and H2O2 were observed between the two isolate groups, S. pseudintermedius isolates exhibited enhanced resistance to PMAP-36 compared to S. schleiferi isolates. CONCLUSIONS: These findings suggest that high a prevalence of various toxin genes together with enhanced resistance to cathelicidins may contribute to the pathogenicity of S. pseudintermedius and S. schleiferi in canine cutaneous infections.

Species Profiles and Antimicrobial Resistance of Non-aureus Staphylococci Isolated from Healthy Broilers, Farm Environments, and Farm Workers.

Non-aureus staphylococci (NAS), particularly antimicrobial-resistant NAS, have a substantial impact on human and animal health. In the current study, we investigated (1) the species profiles of NAS isolates collected from healthy broilers, farm environments, and farm workers in Korea, (2) the occurrence of antimicrobial-resistant NAS isolates, especially methicillin resistance, and (3) the genetic factors involved in the methicillin and fluoroquinolone resistance. In total, 216 NAS isolates of 16 different species were collected from healthy broilers (n=178), broiler farm environments (n=18), and farm workers (n=20) of 20 different broiler farms. The two most dominant broiler-associated NAS species were Staphylococcus agnetis (23.6%) and Staphylococcus xylosus (22.9%). Six NAS isolates were mecA-positive carrying staphylococcal cassette chromosome mec (SCCmec) II (n=1), SCCmec IV (n=1), SCCmec V (n=2), or non-typeable SCCmec element (n=2). While two mecA-positive Staphylococcus epidermidis isolates from farm workers had SCCmec II and IV, a mecA-positive S. epidermidis isolate from broiler and a Staphylococcus haemolyticus isolate farm environment carried SCCmec V. The occurrence of multidrug resistance was observed in 48.1% (104/216 isolates) of NAS isolates with high resistance rates to ß-lactams (>40%) and fusidic acid (59.7%). Fluoroquinolone resistance was confirmed in 59 NAS isolates (27.3%), and diverse mutations in the quinolone resistance determining regions of gyrA, gyrB, parC, and parE were identified. These findings suggest that NAS in broiler farms may have a potential role in the acquisition, amplification, and transmission of antimicrobial resistance.

Inhibition of Listeria monocytogenes in Fresh Cheese Using a Bacteriocin-Producing Lactococcus lactis CAU2013 Strain.

In recent years, biocontrol of foodborne pathogens has become a concern in the food industry, owing to safety issues. Listeria monocytogenes is one of the foodborne pathogens that causes listeriosis. The major concern in the control of L. monocytogenes is its viability as it can survive in a wide range of environments. The purpose of this study was to isolate lactic acid bacteria with antimicrobial activity, evaluate their applicability as a cheese starter, and evaluate their inhibitory effects on L. monocytogenes. Lactococcus lactis strain with antibacterial activity was isolated from raw milk. The isolated strain was a low acidifier, making it a suitable candidate as an adjunct starter culture. The commercial starter culture TCC-3 was used as a primary starter in this study. Fresh cheese was produced using TCC-3 and L. lactis CAU2013 at a laboratory scale. Growth of L. monocytogenes (5 Log CFU/g) in the cheese inoculated with it was monitored during the storage at 4°C and 10°C for 5 days. The count of L. monocytogenes was 1 Log unit lower in the cheese produced using the lactic acid bacteria strain compared to that in the cheese produced using the commercial starter. The use of bacteriocin-producing lactic acid bacteria as a starter culture efficiently inhibited the growth of L. monocytogenes. Therefore, L. lactis can be used as a protective adjunct starter culture for cheese production and can improve the safety of the product leading to an increase in its shelf-life.

Clonal distribution and antimicrobial resistance of methicillin-susceptible and -resistant Staphylococcus aureus strains isolated from broiler farms, slaughterhouses, and retail chicken meat.

Colonization of food-producing animals by antimicrobial-resistant Staphylococcus aureus, especially methicillin-resistant S. aureus (MRSA), has become a serious public health problem worldwide. In the current study, clonal diversities of livestock-associated S. aureus isolates collected from broiler farms, slaughterhouses, and retail chicken meat were examined. Two-hundred S. aureus isolates (43 MRSA and 157 methicillin-susceptible S. aureus [MSSA] isolates) were analyzed to determine 1) the genotypes of the isolates (multilocus sequence, agr, and spa types), 2) the methicillin resistance phenotype and staphylococcal cassette chromosome mec (SCCmec) types, 3) the antimicrobial resistance profiles, and 4) the mutational changes in gyrA, gyrB, parC, and parE in fluoroquinolone-resistant isolates. Fifteen different sequence types (STs) of MSSA strains displaying a relatively high degree of genetic diversity were detected in broiler farms, slaughterhouses, and retail chicken meat. In contrast to MSSA, 2 dominant genetic lineages of MRSA (ST692-SCCmecV with t2249 spa type, and ST188-SCCmecIVa with spa type t189) were found in healthy broilers. The high prevalence of ST692 and ST188 in healthy broilers is associated with high levels of multiple antimicrobial-resistance phenotypes, particularly fluoroquinolone resistance. All fluoroquinolone-resistant isolates carried double point mutations in gyrA (S84L) and parC (S80F), regardless of STs or methicillin resistance. Notably, only the ST188 lineage carried an additional third mutation in gyrB (D494N), correlating with enhanced ciprofloxacin minimum inhibitory concentration values versus the strains with double mutations. These results provide important insights into the genetic diversity of antimicrobial-resistant S. aureus strains associated with the chicken meat production chain, including healthy broilers, in Korea.

Multilocus sequence type-dependent activity of human and animal cathelicidins against community-, hospital-, and livestock-associated methicillin-resistant Staphylococcus aureus isolates.

Sequence type (ST) 5 methicillin-resistant Staphylococcus aureus (MRSA) with staphylococcal cassette chromosome mec (SCCmec) type II (ST5-MRSA-II) and ST72-MRSA-IV represent the most significant genotypes for healthcare- (HA) and community-associated (CA) MRSA in Korea, respectively. In addition to the human-type MRSA strains, the prevalence of livestock-associated (LA) MRSA clonal lineages, such as ST541 and ST398 LA-MRSA-V in pigs and ST692 LA-MRSA-V and ST188 LA-MRSA-IV in chickens, has recently been found. In this study, clonotype-specific resistance profiles to cathelicidins derived from humans (LL-37), pigs (PMAP-36), and chickens (CATH-2) were examined using six different ST groups of MRSA strains: ST5 HA-MRSA-II, ST72 CA-MRSA-IV, ST398 LA-MRSA-V, ST541 LA-MRSA-V, ST188 LA-MRSA-IV, and ST692 LA-MRSA-V. Phenotypic characteristics often involved in cathelicidin resistance, such as net surface positive charge, carotenoid production, and hydrogen peroxide susceptibility were also determined in the MRSA strains. Human- and animal-type MRSA strains exhibited clonotype-specific resistance profiles to LL-37, PMAP-36, or CATH-2, indicating the potential role of cathelicidin resistance in the adaptation and colonization of human and animal hosts. The ST5 HA-MRSA isolates showed enhanced resistance to all three cathelicidins and hydrogen peroxide than ST72 CA-MRSA isolates by implementing increased surface positive charge and carotenoid production. In contrast, LA-MRSA strains employed mechanisms independent of surface charge regulation and carotenoid production for cathelicidin resistance. These results suggest that human- and livestock-derived MRSA strains use different strategies to counteract the bactericidal action of cathelicidins during the colonization of their respective host species.

Profiles of Non-aureus Staphylococci in Retail Pork and Slaughterhouse Carcasses: Prevalence, Antimicrobial Resistance, and Genetic Determinant of Fusidic Acid Resistance.

As commensal colonizers in livestock, there has been little attention on staphylococci, especially non-aureus staphylococci (NAS), contaminating meat production chain. To assess prevalence of staphylococci in retail pork and slaughterhouse carcass samples in Korea, we collected 578 samples from Korean slaughterhouses (n=311) and retail markets (n=267) for isolation of staphylococci and determined antimicrobial resistance phenotypes in all the isolates. The presence of and prevalence of fusB-family genes (fusB, fusC, fusD, and fusF) and mutations in fusA genes were examined in fusidic acid resistant isolates. A total of 47 staphylococcal isolates of 4 different species (Staphylococcus aureus, n=4; S. hyicus, n=1; S. epidermidis, n=10; Mammaliicoccus sciuri, n=32) were isolated. Fusidic acid resistance were confirmed in 9/10 S. epidermidis and all of the 32 M. sciuri (previously S. sciuri) isolates. Acquired fusidic acid resistance genes were detected in all the resistant strains; fusB and fusC in S. epidermidis and fusB/C in M. sciuri. Multi-locus sequence type analysis revealed that ST63 (n=10, 31%) and ST30 (n=8, 25%) genotypes were most prevalent among fusidic acid resistant M. sciuri isolates. In conclusion, the high prevalence of fusB-family genes in S. epidermidis and M. sciuri strains isolated from pork indicated that NAS might act as a reservoir for fusidic acid resistance gene transmissions in pork production chains.

Co-occurrence of cfr-mediated linezolid-resistance in ST398 LA-MRSA and non-aureus staphylococci isolated from a pig farm.

Linezolid resistance, mediated by the cfr gene, which confers resistant phenotypes to phenicols, lincosamides, oxazolidinones, pleuromutilins, and streptogramin A antimicrobials, has emerged in S. aureus and non-aureus staphylococci (NAS). Moreover, due to the transferable potential via plasmids, the spread of cfr among staphylococci is of great concern. In the present study, we investigated the prevalence of cfr-mediated linezolid resistance in ST398 methicillin-resistant S. aureus (MRSA) and NAS strains isolated from a pig farm. Among the 26 staphylococci isolates collected from a pig farm, 14 cfr-harboring ST398 MRSA and NAS (S. epidermidis, S. pasteuri, S. cohnii, and S. rostri) strains were resistant to linezolid and also carried the fexA gene. Comparative genome analysis of cfr-carrying linezolid-resistant ST398 MRSA and NAS (S. pasteuri, S. cohnii, and S. epidermidis) strains revealed that the segments harboring cfr in different staphylococcal strains showed ≥ 99 % sequence identity and the corresponding region containing the cfr, fexA, and Tn558 elements were located in a 38-kb plasmid, designated pSA12 of ST398 MRSA. These observations indicate that the cfr-carrying plasmids and/or fragments may be disseminated among staphylococci in a pig farm and possibly transmitted to staphylococci of human origin, subsequently posing a threat to public health. This is the first report of the co-existence of cfr in linezolid-resistant ST398 MRSA and NAS isolated from a pig farm in South Korea.

Whey fermented by Enterococcus faecalis M157 exhibits antiinflammatory and antibiofilm activities against oral pathogenic bacteria.

The aim of the present study was to investigate the antiinflammatory and antibiofilm effects of whey fermented by Enterococcus faecalis M157 (M157-W) against oral pathogenic bacteria. The M157-W significantly inhibited IL-1ß, IL-6, and nitric oxide induced by the lipopolysaccharide of Porphyromonas gingivalis in RAW 264.7 cells. The M157-W also inhibited the production of IL-1ß and IL-8 in human periodontal ligament cells. Treatment with M157-W suppressed the phosphorylation of mitogen-activated protein kinases as well as the activation of nuclear factor-κB in RAW 264.7 cells stimulated by P. gingivalis lipopolysaccharide. Furthermore, M157-W dose-dependently inhibited Streptococcus mutans biofilm, whereas unfermented whey did not inhibit the biofilm. Treatment with M157-W significantly suppressed gtfB, gtfC, and gtfD gene expression in S. mutans compared with the control (0 µg/mL), indicating that M157-W inhibits S. mutans biofilm formation by reducing the synthesis of extracellular polymeric substances. Collectively, these results suggest that M157-W has antiinflammatory and antibiofilm activities against oral pathogenic bacteria.

Complete genome sequence of Clostridium perfringens B20, a bacteriocin-producing pathogen.

Clostridium perfringens B20 was isolated from chicken feces collected from a local farm associated with Chung-Ang University (Anseong, Korea). The whole genome of C. perfringens B20 was sequenced using the PacBio RS II platform and assembled de novo. The genome is 2,982,563 bp long and assembled in two contigs. Annotation analyses revealed 2,668 protein-coding sequences, 30 rRNA genes, and 94 tRNA genes, with 28.2% G + C (guanine + cytosine) content. In silico genomic analysis revealed the presence of genes encoding a class IId bacteriocin, lactococcin A, and associated ABC transporter and immunity proteins, as well as a putative bacteriocin gene.

Differentiation of carbon black from black carbon using a ternary plot based on elemental analysis.

Carbon black (CB) is composed of engineered nanoparticles that are commercially produced by partial combustion of hydrocarbons. It is mainly used as a reinforcing agent in vehicle tires. Although the potential health effects of CB have been investigated extensively, some toxicological reports interchange CB with black carbon (BC), which has similar features, thereby misusing the term. BC is an undesirable byproduct of the incomplete combustion of fuels. Therefore, there is a need to differentiate CB from the unintentionally produced nanomaterials (BC) in nano-toxicity, environmental, and human health studies. To distinguish clearly CB from BC, it is important to find the key parameters from several characteristics of two substances. The fundamental physicochemical properties of commercial CB and naturally formed BC were conducted. Based on the elemental analysis, we found three key factors, which could be used to differentiate the CB from BC. And thus, herein, we propose a ternary plot of the aH/C-log(C/b)-1/H combination for use in differentiating CB from BC. The plot of the 100H/C-log(C/10)-1/H combination of elemental ratios separated the CB domain from the BC domain symmetrically. The effectiveness of the ternary chart was validated using 37 samples (nine samples in this work, 25 sample results taken from references studies, and three samples from the field). Therefore, the ternary plot could be used as a prescreening tool for distinguishing CB from BC.

Chryseobacterium vaccae sp. nov., isolated from raw cow's milk.

Strain CA7T, a Gram-stain-negative, non-motile, non-spore-forming, aerobic and rod-shaped bacterial strain, was isolated from raw cow's milk collected from a farm affiliated with Chung-Ang University, Anseong, Korea, and characterized by a polyphasic approach. Optimal growth of strain CA7T was observed on tryptic soy agar at 30 °C and pH 7.0 with 0 % NaCl. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that strain CA7T belonged to the genus Chryseobacterium. The most closely related strains (16S rRNA gene sequence similarity indicated in parentheses), based on the phylogenetic analysis, were Chryseobacterium rhizosphaerae KCTC 22548T (98.08 %), Chryseobacterium nakagawai CCUG 60563T (98.61 %), Chryseobacterium jejuense KACC 12501T (97.85 %) and Chryseobacterium aurantiacum KCTC 62135T (97.78 %). Whole genome sequencing indicated that the genome size was 5 125 723 bp and had a DNA G+C content of 37.4 mol%. Average nucleotide identity values for strain CA7T with C. rhizosphaerae, C. nakagawai, C. jejuense, C. aurantiacum, and the type species of the genus Chryseobacterium, C. gleum, were 80.2, 79.8, 79.8, 79.6 and 80.4 %, respectively. The digital DNA-DNA hybridization values of CA7T compared to C. rhizosphaerae, C. nakagawai, C. jejuense, C. aurantiacum and C. gleum were 24.1, 23.9, 23.9, 23.7 and 24.3 %, respectively. The major fatty acids were iso-C15 : 0, summed feature 9 (iso-C17 : 1 ω9c and/or C16 : 0 10-methyl), iso-C17 : 0 3-OH and summed feature 3 (iso-C15 : 0 2-OH and/or C16 : 1 ω7c). Menaquinone-6 was the only respiratory quinone. The major polar lipid was phosphatidylethanolamine. Based on this polyphasic taxonomic study, strain CA7T represents a novel species of the genus Chryseobacterium for which the name Chryseobacterium vaccae sp. nov. is proposed. The type strain is CA7T (=KACC 21402T=JCM 33749T).

Complete genome sequence of Streptococcus hyointestinalis B19, a strain producing bacteriocin, isolated from chicken feces.

Streptococcus hyointestinalis B19 was isolated from chicken feces collected from local farm in Anseong, Korea. S. hyointestinalis B19 was shown to produce bacteriocin-like compounds exhibiting inhibitory activities against several pathogens including strains of Clostridium perfringens and Listeria monocytogenes. The whole genome of S. hyointestinalis B19 strain was sequenced using PacBio RS II platform. The genome comprised four contigs with a size of 2,217,061 bp. The DNA G + C content was found to be 42.95 mol%. Annotation results revealed 2,266 coding sequences (CDSs), 18 rRNAs, and 61 tRNA genes. Based on genome analysis, we found that the strain B19 possessed various genes associated with bacteriocin synthesis, modification, and transport.

Acinetobacter pullorum sp. nov., Isolated from Chicken Meat.

A bacterial strain, designated B301T and isolated from raw chicken meat obtained from a local market in Korea, was characterized and identified using a polyphasic taxonomic approach. Cells were gram-negative, non-motile, obligate-aerobic coccobacilli that were catalase-positive and oxidase-negative. The optimum growth conditions were 30°C, pH 7.0, and 0% NaCl in tryptic soy broth. Colonies were round, convex, smooth, and cream-colored on tryptic soy agar. Strain B301T has a genome size of 3,102,684 bp, with 2,840 protein-coding genes and 102 RNA genes. The 16S rRNA gene analysis revealed that strain B301T belongs to the genus Acinetobacter and shares highest sequence similarity (97.12%) with A. celticus ANC 4603T and A. sichuanensis WCHAc060041T. The average nucleotide identity and digital DNA-DNA hybridization values for closely related species were below the cutoff values for species delineation (95-96% and 70%, respectively). The DNA G+C content of strain B301T was 37.0%. The major respiratory quinone was Q-9, and the cellular fatty acids were primarily summed feature 3 (C16:1 ω6c/C16:1 ω7c), C16:0, and C18:1 ω9c. The major polar lipids were phosphatidylethanolamine, diphosphatidyl-glycerol, phosphatidylglycerol, and phosphatidyl-serine. The antimicrobial resistance profile of strain B301T revealed the absence of antibiotic-resistance genes. Susceptibility to a wide range of antimicrobials, including imipenem, minocycline, ampicillin, and tetracycline, was also observed. The results of the phenotypic, chemotaxonomic, and phylogenetic analyses indicate that strain B301T represents a novel species of the genus Acinetobacter, for which the name Acinetobacter pullorum sp. nov. is proposed. The type strain is B301T (=KACC 21653T = JCM 33942T).

Acinetobacter pullicarnis sp. nov. isolated from chicken meat.

A novel bacterial strain, named S23T, was isolated from chicken meat of local market in Korea. Cells were Gram-negative, milky-yellow colored, non-motile and coccobacillus. The strain was obligate aerobic and catalase-positive, oxidase-negative, optimum growth temperature and pH were 25 °C and pH 7.0, respectively. On the basis of 16S rRNA gene sequence analysis, strain S23T belongs to the genus Acinetobacter and is most closely related to Acinetobacter defluvii KCTC 52503 T (97.40%). The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) value between strain S23T and its closet phylogenetic neighbors was below 76% and 17%, respectively. The G + C content of genomic DNA of strain S23T was 41.53 mol%. The major respiratory quinone was Q-9. The major cellular fatty acids were summed feature 3 (comprising C16:1ω7c and/or C16:1ω6c), C18:1ω9c, and C16:0. The major polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanol-amine, and phosphatidylserine. The ANI and dDDH results and results of the genotypic analysis in combination with chemotaxonomic and physiological data demonstrated that strain S23T represented a novel species within the genus Acinetobacter, for which the name Acinetobacter pullicarnis sp. nov. is proposed. The strain type is S23T (= KACC 19921 T = JCM 33150 T).

Chryseobacterium mulctrae sp. nov., isolated from raw cow's milk.

A Gram-stain-negative bacterial strain, designated CA10T, was isolated from bovine raw milk sampled in Anseong, Republic of Korea. Cells were yellow-pigmented, aerobic, non-motile bacilli and grew optimally at 30 °C and pH 7.0 on tryptic soy agar without supplementation of NaCl. Phylogenetic analysis based on the 16S rRNA gene sequences revealed that strain CA10T belonged to the genus Chryseobacterium, family Flavobacteriaceae, and was most closely related to Chryseobacterium indoltheticum ATCC 27950T (98.75 % similarity). The average nucleotide identity and digital DNA-DNA hybridization values of strain CA10T were 94.4 and 56.9 %, respectively, relative to Chryseobacterium scophthalmum DSM 16779T, being lower than the cut-off values of 95-96 and 70 %, respectively. The predominant respiratory quinone was menaquinone-6; major polar lipid, phosphatidylethanolamine; major fatty acids, iso-C15 : 0, summed feature 9 (iso-C17 : 1ω9c and/or C16 : 0 10-methyl), summed feature 3 (iso-C15 : 0 2-OH and/or C16 : 1ω7c) and iso-C17 : 0 3-OH. The results of physiological, chemotaxonomic and biochemical analyses suggested that strain CA10T is a novel species of genus Chryseobacterium, for which the name Chryseobacterium mulctrae sp. nov. is proposed. The type strain is CA10T (=KACC 21234T=JCM 33443T).

Lactobacillus plantarum CAU1055 ameliorates inflammation in lipopolysaccharide-induced RAW264.7 cells and a dextran sulfate sodium-induced colitis animal model.

This study aimed to screen lactic acid bacteria (LAB) for their anti-inflammatory activity by using RAW264.7 cells and dextran sulfate sodium (DSS)-induced colitis. In all, 192 LAB strains were isolated from healthy human feces, of which 8 strains showed excellent nitric oxide (NO) inhibitory activity. Peptidoglycan extracts of these 8 LAB strains were subjected to NO assay, Western blot, and ELISA. Among the 8 tested strains, extracts of 4 strains significantly inhibited the production of NO, related enzyme activities such as inducible nitric oxide synthase and cyclooxygenase 2, and key cytokines such as tumor necrosis factor-α and IL-6 in RAW264.7 cells. The 4 strains belonged to Lactobacillus (CAU1054, CAU1055, CAU1064, and CAU1301). Oral administration of the 4 strains inhibited DSS-induced body weight loss, colon shortening, and colon damage in ICR mice. The colon tissue of the mice treated with Lactobacillus plantarum strain CAU1055 had significantly reduced levels of inducible nitric oxide synthase, cyclooxygenase 2, tumor necrosis factor-α, and IL-6. We found that strain CAU1055 could be used as a candidate probiotic strain for the prevention and treatment of inflammatory bowel disease. Further studies are warranted to confirm the mechanisms of interaction between peptidoglycan of L. plantarum strain CAU1055 and upstream cellular signaling mediators.

Chryseobacterium aureum sp. nov., isolated from the Han River, Republic of Korea.

A Gram-stain-negative, yellow-pigmented, non-motile, non-spore-forming, aerobic and rod-shaped bacterial strain, designated 17S1E7T, was isolated from the Han River, Republic of Korea, and characterized by polyphasic taxonomy analyses. Strain 17S1E7T grew optimally on tryptic soy agar at 37 °C and pH 7.0 in the absence of NaCl. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain 17S1E7T belonged to the genus Chryseobacterium and was most closely related to Chryseobacterium culicis DSM 23031T (98.54 %). The average nucleotide identity value of strain 17S1E7T was 91.1 % to Chryseobacterium culicis DSM 23031T, which was lower than the cut-off of 95-96 %. The DNA G+C content of strain 17S1E7T was 37.4 mol%. Flexirubin-type pigments were produced. The predominant respiratory quinone was menaquinone 6. The major fatty acids of strain 17S1E7T were iso-C15 : 0, summed feature 9 (iso-C17 : 1ω9c and/or C16 : 0 10-methyl), iso-C17 : 0 3-OH and summed feature 3 (iso-C15 : 0 2-OH and/or C16 : 1ω7c). The predominant polar lipid was phosphatidylethanolamine. Based on polyphasic taxonomy data, strain 17S1E7T represents a novel species of the genus Chryseobacterium, for which the name Chryseobacterium aureum sp. nov. is proposed. The type strain is 17S1E7T (=KACC 19920T=JCM 33165T).

Identification of duck liver-expressed antimicrobial peptide 2 and characterization of its bactericidal activity.

OBJECTIVE: This study was conducted to identify duck liver-expressed antimicrobial peptide 2 (LEAP-2) and demonstrate its antimicrobial activity against various pathogens. METHODS: Tissue samples were collected from 6 to 8-week-old Pekin ducks (Anas platyrhynchos domesticus), total RNA was extracted, and cDNA was synthesized. To confirm the duck LEAP-2 transcript expression levels, quantitative real-time polymerase chain reaction was conducted. Two kinds of peptides (a linear peptide and a disulfide-type peptide) were synthesized to compare the antimicrobial activity. Then, antimicrobial activity assay and fluorescence microscopic analysis were conducted to demonstrate duck LEAP-2 bactericidal activity. RESULTS: The duck LEAP-2 peptide sequence showed high identity with those of other avian species (>85%), as well as more than 55% of identity with mammalian sequences. LEAP-2 mRNA was highly expressed in the liver with duodenum next, and then followed by lung, spleen, bursa and jejunum and was the lowest in the muscle. Both of LEAP-2 peptides efficiently killed bacteria, although the disulfide-type LEAP-2 showed more powerful bactericidal activity. Also, gram-positive bacteria was more susceptible to duck LEAP-2 than gram-negative bacteria. Using microscopy, we confirmed that LEAP-2 peptides could kill bacteria by disrupting the bacterial cell envelope. CONCLUSION: Duck LEAP-2 showed its antimicrobial activity against both gram-positive and gram-negative bacteria. Disulfide bonds were important for the powerful killing effect by disrupting the bacterial cell envelope. Therefore, duck LEAP-2 can be used for effective antibiotics alternatives.