Pathogenic Potential and Control of Chryseobacterium Species from Clinical, Fish, Food and Environmental Sources.

Chryseobacterium species are isolated and taxonomically evaluated from a wide range of sources. While C. gleum and C. indologenes have been implicated in human disease, the potential pathogenicity of numerous other species have not been investigated. The aims were therefore to evaluate 37 Chryseobacterium species and Elizabethkingia meningoseptica from environmental, food, fish, water and clinical sources for production of haemolysis, growth at 37 °C, and production of virulence enzymes. The control of these strains were investigated by determination of antimicrobial and disinfectant resistance. All the species produced α- or ß-haemolysis. In terms of growth at 37 °C and production of virulence enzymes, C. soldanellicola (environmental), C. oranimense (food) and C. koreense (natural mineral water) could be potential human pathogens. Chryseobacterium piscium might be pathogenic to fish. Trimethoprim could be the most effective antimicrobial for the treatment of a Chryseobacterium species infection, while the disinfectants that contain poly-dimethyl ammonium chloride or benzalkonium chloride could be regarded as the most effective for decontamination of surfaces contaminated with Chryseobacterium species.

Chryseobacterium pennae sp. nov., isolated from poultry feather waste.

A Gram-stain-negative, rod-shaped, non-motile, non-spore-forming, aerobic, yellow-pigmented bacterium was isolated from chicken feather waste collected from an abattoir in Bloemfontein, South Africa. A polyphasic taxonomy study was used to describe and name the bacterial isolate, strain 1_F178T. The 16S rRNA gene sequence analysis and sequence comparison data indicated that strain 1_F178T was a member of the genus Chryseobacterium and was closely related to Chryseobacterium jejuense (99.1%) and Chryseobacterium nakagawai (98.7%). Overall genome similarity metrics (average nucleotide identity, digital DNA-DNA hybridization and average amino acid identity) revealed greatest similarity to the C. jejuense and C. nakagawai type strains but were below the threshold for species delineation. Genome sequencing revealed a genome size of 6.18 Mbp and a G+C content of 35.6 mol%. The major respiratory quinone and most abundant polar lipid of strain 1_F178T were menaquinone-6 and phosphatidylethanolamine, respectively. Strain 1_F178T had a typical fatty acid composition for Chryseobacterium species. On the basis of physiological, genotypic, phylogenetic and chemotaxonomic data, strain 1_F178T constitutes a novel species of Chryseobacterium, for which the name Chryseobacterium pennae sp. nov. is proposed. The type strain is 1_F178T (=LMG 30779T=KCTC 62759T).

Heterologous expression and characterisation of a keratinase produced by Chryseobacterium carnipullorum.

Chryseobacterium carnipullorum 9_R23581T, isolated from raw chicken meat, was evaluated for its potential to degrade keratin found in feathers. The focus of this study was to heterologously express and characterise a keratinolytic enzyme produced by C. carnipullorum. Chryseobacterium carnipullorum secretes proteolytic enzymes that have feather degrading capabilities during its exponential growth phase. This study concluded that the most likely main component of the keratinolytic enzymes of C. carnipullorum was peptidase M64, a serine-endopeptidase with a molecular weight in crude form of 49.46 kDa. Primers were designed on the selected gene of interest, which was amplified from the genome of C. carnipullorum (accession number NZ-FRCD01000002.1). The gene coding for peptidase M64 was further cloned, propagated and expressed in E. coli BL21 [DE3] cells. Purification was by Immobilised Metal Affinity Chromatography (IMAC). The molecular weight of the keratinase was about 50 kDa after purification while its optimum temperature and pH were 50 °C and 8.5, respectively. The activity of this keratinase was inhibited by phenylmethylsulfonyl fluoride (PMSF) and it was enhanced by the presence of divalent metal ions such as Mg2+ and Ca2+. Enzyme activity was further assayed by application to chicken feathers and observed degradation was an indication of keratinolytic potential.

Division of the genus Chryseobacterium: Observation of discontinuities in amino acid identity values, a possible consequence of major extinction events, guides transfer of nine species to the genus Epilithonimonas, eleven species to the genus Kaistella, and three species to the genus Halpernia gen. nov., with description of Kaistella daneshvariae sp. nov. and Epilithonimonas vandammei sp. nov. derived from clinical specimens.

The genus Chryseobacterium in the family Weeksellaceae is known to be polyphyletic. Amino acid identity (AAI) values were calculated from whole-genome sequences of species of the genus Chryseobacterium, and their distribution was found to be multi-modal. These naturally-occurring non-continuities were leveraged to standardise genus assignment of these species. We speculate that this multi-modal distribution is a consequence of loss of biodiversity during major extinction events, leading to the concept that a bacterial genus corresponds to a set of species that diversified since the Permian extinction. Transfer of nine species (Chryseobacterium arachidiradicis, Chryseobacterium bovis, Chryseobacterium caeni, Chryseobacterium hispanicum, Chryseobacterium hominis, Chryseobacterium hungaricum,, Chryseobacterium pallidum and Chryseobacterium zeae) to the genus Epilithonimonas and eleven (Chryseobacterium anthropi, Chryseobacterium antarcticum, Chryseobacterium carnis, Chryseobacterium chaponense, Chryseobacterium haifense, Chryseobacterium jeonii, Chryseobacterium montanum, Chryseobacterium palustre, Chryseobacterium solincola, Chryseobacterium treverense and Chryseobacterium yonginense) to the genus Kaistella is proposed. Two novel species are described: Kaistella daneshvariae sp. nov. and Epilithonimonas vandammei sp. nov. Evidence is presented to support the assignment of Planobacterium taklimakanense to a genus apart from Chryseobacterium, to which Planobacterium salipaludis comb nov. also belongs. The novel genus Halpernia is proposed, to contain the type species Halpernia frigidisoli comb. nov., along with Halpernia humi comb. nov., and Halpernia marina comb. nov.

Chryseobacterium pennipullorum sp. nov., isolated from poultry feather waste.

Strain 7_F195T was previously isolated from chicken feather waste collected from an abattoir in Bloemfontein, South Africa. A polyphasic approach was followed to determine if strain 7_F195T belongs to the genus Chryseobacterium and if the organism can be classified as a new species. The nearest neighbours, based on 16S rRNA gene sequence similarity values (indicated in parentheses), were Chryseobacterium flavum KCTC 12877T (98.42 %), Chryseobacterium indologenesLMG 8337T (98.24 %) and Chryseobacterium gleum ATCC 35910T (97.71 %). Genome sequencing revealed a genome size of 4 796 535 bp and a DNA G+C content of 38.6 mol%. The ANI values of strain 7_F195T compared to C. flavum, C. indologenesand C. gleum were 81.45, 81.86 and 82.38 %, respectively. The digital DNA-DNA hybridization values for strain 7_F195T with C. flavum, C. indologenes and C. gleum were 23.7, 23.7 and 24.9 %, respectively. Notable phenotypic differences include the presence of urease activity in C. indologenes LMG 8337T and C. gleum NCTC 11432T, but not in strain 7_F195T or C. flavum KCTC 12877T. The predominant fatty acids of strain 7_F195T were iso-C15 : 0, iso-C17 : 1ω9c and iso-C17 : 0 3-OH and the most abundant polar lipid was phosphatidylethanolamine. Menaquinone-6 was the only respiratory quinone. Based on the data generated from this polyphasic study, strain 7_F195T represents a novel Chryseobacterium species for which the name Chryseobacteriumpennipullorum sp. nov. is proposed. The type strain is 7_F195T (=LMG 30781T=KCTC 62760T).

Draft Genome Sequences of Seven Chryseobacterium Type Strains.

In an honors course on "Omics Sciences," draft genome sequences of Chryseobacterium elymi KCTC 22547T, Chryseobacterium flavum KCTC 12877T, Chryseobacterium hispanicum KCTC 22104T, Chryseobacterium lathyri KCTC 22544T, "Candidatus Chryseobacterium massiliae" CCUG 51329T, Chryseobacterium piscium CCUG 51923T, and Chryseobacterium rhizosphaerae KCTC 22548T were generated to facilitate phylogenomic comparisons within the genus.

The chemical, microbial, sensory and technological effects of intermediate salt levels as a sodium reduction strategy in fresh pork sausages.

BACKGROUND: The reduction of sodium in processed meat products is synonymous with the use of salt replacers. Rarely has there been an assessment of the use of intermediate salt levels as a sodium reduction strategy in itself. In this study, 1 and 1.5% salt levels were compared with 0 and 2% controls in fresh pork sausages for effects on chemical, microbial, sensory and technological stability. RESULTS: Although significant (P < 0.001 to P < 0.01) differences were found between the 0 and 2% controls, no significant differences could be detected between the 2, 1.5 and 1% added NaCl treatments for the following: total bacteria counts on days 3, 6 and 9; TBARS of pork sausages stored at 4 °C on days 6 and 9 and stored at -18 °C on days 90 and 180; taste, texture and overall liking during sensory evaluation; and % cooking loss, % total loss and % refrigeration loss. Consumers were able to differentiate between the 2 and 1% added NaCl treatments in terms of saltiness. CONCLUSION: This study indicated that salt reduction to intermediate levels can be considered a sodium reduction strategy in itself but that further research with regards to product safety is needed. © 2015 Society of Chemical Industry.

Characterization of bacterial pathogens in rural and urban irrigation water.

The study aimed to compare the bacteriological quality of an urban and rural irrigation water source. Bacterial counts, characterization, identification and diversity of aerobic bacteria were determined. Escherichia coli isolated from both sites was subjected to antibiotic susceptibility testing, virulence gene (Stx1/Stx2 and eae) determination and (GTG)5 Rep-PCR fingerprinting. Low mean monthly counts for aerobic spore formers, anaerobic spore formers and Staphylococcus aureus were noted although occasional spikes were observed. The most prevalent bacterial species at both sites were Bacillus spp., E. coli and Enterobacter spp. In addition, E. coli and Bacillus spp. were most prevalent in winter and summer respectively. Resistance to at least one antibiotic was 84% (rural) and 83% (urban). Highest resistance at both sites was to cephalothin and ampicillin. Prevalence of E. coli possessing at least one virulence gene (Stx1/Stx2 and eae) was 15% (rural) and 42% (urban). All (rural) and 80% (urban) of E. coli possessing virulence genes showed antibiotic resistance. Complete genetic relatedness (100%) was shown by 47% of rural and 67% of urban E. coli isolates. Results from this study show that surface irrigation water sources regardless of geographical location and surrounding land-use practices can be reservoirs of similar bacterial pathogens.

Chryseobacterium carnipullorum sp. nov., isolated from raw chicken.

Three Gram-staining-negative, rod-shaped, non-spore-forming, non-motile, oxidase-positive, yellow pigmented and aerobic bacterial isolates designated 8_R23573, 9_R23581(T) and 10_R23577 were isolated from raw chicken at a broiler processing plant in Bloemfontein, South Africa. A polyphasic taxonomic approach was used to determine their exact taxonomic identities. Phylogenetic analysis of the 16S rRNA gene sequences showed that the three strains belonged to the genus Chryseobacterium, exhibiting the highest similarities to Chryseobacterium shigense DSM 17126(T) (98.6-99.2%) and Chryseobacterium luteum DSM 18605(T) (98.3-98.7%). The most abundant quinone was menaquinone MK-6 and the predominant cellular fatty acids were iso-15:0, iso-17:1ω9c, iso-17:0 3-OH and summed feature 3 (iso-16:1ω7c and/or iso-15:0 2-OH), which supported the affiliation of the strains to the genus Chryseobacterium. The DNA G+C contents of the strains were 36.9, 36.7 and 36.6 mol% respectively. The DNA-DNA hybridization results gave relatedness values ranging from 78.8 to 87.2% among the three strains and 23.4 to 56.1% to the two nearest phylogenetic neighbours C. shigense DSM 17126(T) and C. luteum LMG 23785(T). On the basis of the data from this polyphasic study, the three strains are concluded to represent a novel species of the genus Chryseobacterium for which the name Chryseobacterium carnipullorum sp. nov. is proposed. The type strain is 9_R23581(T) ( = LMG 26732(T) =DSM 25581(T)).

Chryseobacterium piscium sp. nov., isolated from fish of the South Atlantic Ocean off South Africa.

Four isolates from freshly caught fish samples obtained from the South Atlantic Ocean off the South African coastline were shown to represent a novel species in the genus Chryseobacterium by means of a polyphasic taxonomic study. The four isolates had virtually identical whole-cell protein profiles, fatty acid profiles and biochemical properties. Analysis of the 16S rRNA sequence of strain LMG 23089(T) revealed 99.3 and 98.9 % similarity to the 16S rRNA sequences of the type strains of Chryseobacterium balustinum and Chryseobacterium scophthalmum, respectively. Strain LMG 23089(T) and the C. balustinum and C. scophthalmum type strains formed a stable lineage supported by a bootstrap value of 100 %. The levels of DNA-DNA hybridization towards these nearest phylogenetic neighbours were below 57 %. The absence of growth on MacConkey agar or at 37 degrees C (on nutrient agar), the capacity to grow in the presence of 5 % NaCl and the production of urease activity differentiate this novel taxon from C. balustinum and C. scophthalmum. The four isolates are formally classified as Chryseobacterium piscium sp. nov., with strain LMG 23089(T) (=CCUG 51923(T)) as the type strain. Its DNA G + C content is 33.6 mol%.

Chryseobacterium vrystaatense sp. nov., isolated from raw chicken in a chicken-processing plant.

Yellow-pigmented, Gram-negative organisms isolated from raw chicken were investigated by means of a polyphasic taxonomic approach and were shown to represent a novel species in the genus Chryseobacterium, for which the name Chryseobacterium vrystaatense sp. nov. is proposed. Its nearest phylogenetic neighbours were Chryseobacterium joostei, Chryseobacterium indologenes and Chryseobacterium gleum, which showed 16S rRNA gene sequence similarity levels of 96.9, 97.1 and 96.1%, respectively. Levels of DNA-DNA hybridization between strains of C. vrystaatense and Chryseobacterium reference species were below 46%. Strain LMG 22846(T) (=CCUG 50970(T)) was chosen as the type strain and has a DNA G+C content of 37.1 mol%.

Chryseobacterium joostei sp. nov., isolated from the dairy environment.

Among a large collection of South African dairy isolates, a novel Chryseobacterium taxon (DNA group 3) was previously delineated by a polyphasic taxonomic study (Hugo et al., Syst Appl Microbiol 22, 586-595, 1999). In the present paper, this taxon is further characterized using 16S rDNA sequencing, fatty acid methyl ester analysis and a comparative phenotypic analysis, resulting in the proposal of a novel species, Chryseobacterium joostei sp. nov. (type strain Ix 5a(T) = LMG 18212(T) = CCUG 46665(T)).