Identification and Characterization of Leuconostoc carnosum, Associated with Production and Spoilage of Vacuum-Packaged, Sliced, Cooked Ham

Leuconostoc carnosum was shown to be the specific spoilage organism in vacuum-packaged, sliced, cooked ham showing spoilage during 3 weeks of shelf life. Identification of the specific spoilage organism was done by use of phenotypic data and ClaI, EcoRI, and HindIII reference strain ribopatterns. One hundred L. carnosum isolates associated with the production and spoilage of the ham were further characterized by pulsed-field gel electrophoresis (PFGE), together with some meat-associated Leuconostoc species: L. citreum, L. gelidum, L. mesenteroides subsp. dextranicum, and L. mesenteroides subsp. mesenteroides. ApaI and SmaI digests divided the industrial L. carnosum strains into 25 different PFGE types, ApaI and SmaI types being consistent. Only one specific PFGE type was associated with the spoiled packages. This type also was detected in air and raw-meat mass samples. The spoilage strain did not produce bacteriocins. Only seven isolates belonging to three different PFGE types produced bacteriocins. Similarity analysis of the industrial L. carnosum strains revealed a homogeneous cluster which could be divided into eight subclusters consisting of strains having at most three-fragment differences. The L. carnosum cluster was clearly distinguished from the other meat-associated leuconostoc clusters, with the exception of the L. carnosum type strain. Ribotyping can be very helpful in the identification of L. carnosum, but its discriminatory power is too weak for strain characterization. PFGE provides good discrimination for studies dealing with the properties of homogeneous L. carnosum strains.     

Integration of Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry and Molecular Cloning for the Identification and Functional Characterization of Mobile ortho-Halobenzoate Oxygenase Genes in Pseudomonas aeruginosa Strain JB2

Protein mass spectrometry and molecular cloning techniques were used to identify and characterize mobile o-halobenzoate oxygenase genes in Pseudomonas aeruginosa strain JB2 and Pseudomonas huttiensis strain D1. Proteins induced in strains JB2 and D1 by growth on 2-chlorobenzoate (2-CBa) were extracted from sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels and analyzed by matrix-assisted laser desorption ionization–time of flight mass spectrometry. Two bands gave significant matches to OhbB and OhbA, which have been reported to be the α and β subunits, respectively, of an ortho-1,2-halobenzoate dioxygenase of P. aeruginosa strain 142 (T. V. Tsoi, E. G. Plotnikova, J. R. Cole, W. F. Guerin, M. Bagdasarian, and J. M. Tiedje, Appl. Environ. Microbiol. 65:2151–2162, 1999). PCR and Southern hybridization experiments confirmed that ohbAB were present in strain JB2 and were transferred from strain JB2 to strain D1. While the sequences of ohbA from strains JB2, D1, and 142 were identical, the sequences of ohbB from strains JB2 and D1 were identical to each other but differed slightly from that of strain 142. PCR analyses and Southern hybridization analyses indicated that ohbAB were conserved in strains JB2 and D1 and in strain 142 but that the regions adjoining these genes were divergent. Expression of ohbAB in Escherichia coli resulted in conversion of o-chlorobenzoates to the corresponding (chloro)catechols with the following apparent affinity: 2-CBa ≈ 2,5-dichlorobenzoate > 2,3,5-trichlorobenzoate > 2,4-dichlorobenzoate. The activity of OhbAB_JB2 appeared to differ from that reported for OhbAB₁₄₂ primarily in that a chlorine in the para position posed a greater impediment to catalysis with the former. Hybridization analysis of spontaneous 2-CBa⁻ mutants of strains JB2 and D1 verified that ohbAB were lost along with the genes, suggesting that all of the genes may be contained in the same mobile element. Strains JB2 and 142 originated from California and Russia, respectively. Thus, ohbAB and/or the mobile element on which they are carried may have a global distribution.     

Transformation of Leuconostoc carnosum 4010 and Evidence for Natural Competence of the Organism

Plasmid transformation in Leuconostoc carnosum 4010 was analyzed. A successful transformation protocol for L. carnosum was established by modifying an existing protocol for Lactococcus lactis. Several parameters, including the number of generations that the cells had grown at the time of harvest, glycine concentration, the time of incubation for phenotypic expression, and the electrical field strength, were investigated and proved to have influence on the transformation frequency. Electrocompetence was found to be transient and to peak in the early exponential growth phase. Optimized conditions resulted in transformation frequencies of up to 6.7 × 10⁵ transformants per microgram of plasmid DNA. A total of five plasmids in L. carnosum were successfully introduced and maintained. Interestingly, we discovered that DNA uptake was of a frequency of 3 × 10⁻⁶ to 19 × 10⁻⁶ transformants per CFU in the absence of an applied electrical field. We concluded that L. carnosum is naturally competent.     

Genetic characterisation and heterologous expression of leucocin C, a class IIa bacteriocin from Leuconostoc carnosum 4010

Leuconostoc carnosum 4010 is a protective culture for meat products. It kills the foodborne pathogen Listeria monocytogenes by producing two class IIa (pediocin-like) bacteriocins, leucocin A and leucocin C. The genes for leucocin A production have previously been characterised from Leuconostoc gelidum UAL 187, whereas no genetic studies about leucocin C has been published. Here, we characterised the genes for the production of leucocins A and C in L. carnosum 4010. In this strain, leucocin A and leucocin C operons were localised in different plasmids. Unlike in L. gelidum, leucocin A operon in L. carnosum 4010 only contained the structural and the immunity genes lcaAB without transporter genes lcaECD. On the contrary, leucocin C cluster included two intact operons. Novel genes lecCI encode the leucocin C precursor and the 97-aa immunity protein LecI, respectively. LecI shares 48 % homology with the immunity proteins of sakacin P and listeriocin. Another leucocin C operon lecXTS, encoding an ABC transporter and an accessory protein, was 97 % identical with the leucocin A transporter operon lcaECD of L. gelidum. For heterologous expression of leucocin C in Lactococcus lactis, the mature part of the lecC gene was fused with the signal sequence of usp45 in the secretion vector pLEB690. L. lactis secreted leucocin C efficiently, as shown by large halos on lawns of L. monocytogenes and Leuconostoc mesenteroides indicators. The function of LecI was then demonstrated by expressing the gene lecI in L. monocytogenes. LecI-producing Listeria was less sensitive to leucocin C than the vector strain, thus corroborating the immunity function of LecI.     

Development of a System for Genetic Manipulation of Bartonella bacilliformis

Lack of a system for site-specific genetic manipulation has severely hindered studies on the molecular biology of all Bartonella species. We report the first site-specific mutagenesis and complementation for a Bartonella species. A highly transformable strain of B. bacilliformis, termed JB584, was isolated and found to exhibit a significant increase in transformation efficiency with the broad-host-range plasmid pBBR1MCS-2, relative to wild-type strains. Restriction analyses of genomic preparations with the methylation-sensitive restriction enzymes ClaI and StuI suggest that strain JB584 possesses a dcm methylase mutation that contributes to its enhanced transformability. A suicide plasmid, pUB1, which contains a polylinker, a pMB1 replicon, and a nptI kanamycin resistance cassette, was constructed. An internal 508-bp fragment of the B. bacilliformis flagellin gene (fla) was cloned into pUB1 to generate pUB508, a fla-targeting suicide vector. Introduction of pUB508 into JB584 by electroporation generated eight Kan^r clones of B. bacilliformis. Characterization of one of these strains, termed JB585, indicated that allelic exchange between pUB508 and fla had occurred. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunoblotting, and electron microscopy showed that synthesis of flagellin encoded by fla and secretion/assembly of flagella were abolished. Complementation of fla in trans was accomplished with a pBBR1MCS recombinant containing the entire wild-type fla gene (pBBRFLAG). These data conclusively show that inactivation of fla results in a bald, nonmotile phenotype and that pMB1 and REP replicons make suitable B. bacilliformis suicide and shuttle vectors, respectively. When used in conjunction with the highly transformable strain JB584, this system for site-specific genetic manipulation and complementation provides a new venue for studying the molecular biology of B. bacilliformis.     

Bacteriocin production ofLeuconostoc carnosum LA54A at different combinations of pH and temperature

Summary Leuconostoc carnosum LA54A produces a bacteriocin which is active againstListeria monocytogenes andListeria innocua. The ability ofLc. carnosum to produce the bacteriocin at various combinations of the growth parameters pH and temperature was analyzed. In the case of this strain bacteriocin production seems to be coupled to growth rate. This fact enables the prediction ifLc. carnosum can produce the bacteriocin at a given set of growth parameters, simply by predicting the growth rate of this organism. In addition we have analyzed the growth behavior of the target organismL. innocua WS2258 at the same set of growth parameters.     

Antibiotic Susceptibility Profiles of Dairy Leuconostoc,Analysis of the Genetic Basis of Atypical Resistances and Transfer of Genes In Vitro and in a Food Matrix

In spite of a global concern on the transfer of antibiotic resistances (AR) via the food chain, limited information exists on this issue in species of Leuconostoc and Weissella, adjunct cultures used as aroma producers in fermented foods. In this work, the minimum inhibitory concentration was determined for 16 antibiotics in 34 strains of dairy origin, belonging to Leuconostoc mesenteroides (18), Leuconostoc citreum (11), Leuconostoc lactis (2), Weissella hellenica (2), and Leuconostoc carnosum (1). Atypical resistances were found for kanamycin (17 strains), tetracycline and chloramphenicol (two strains each), and erythromycin, clindamycin, virginiamycin, ciprofloxacin, and rifampicin (one strain each). Surprisingly, L. mesenteroides subsp. mesenteroides LbE16, showed resistance to four antibiotics, kanamycin, streptomycin, tetracycline and virginiamycin. PCR analysis identified tet(S) as responsible for tetracycline resistance in LbE16, but no gene was detected in a second tetracycline-resistant strain, L. mesenteroides subsp. cremoris LbT16. In Leuconostoc mesenteroides subsp. dextranicum LbE15, erythromycin and clindamycin resistant, an erm(B) gene was amplified. Hybridization experiments proved erm(B) and tet(S) to be associated to a plasmid of ≈35 kbp and to the chromosome of LbE15 and LbE16, respectively. The complete genome sequence of LbE15 and LbE16 was used to get further insights on the makeup and genetic organization of AR genes. Genome analysis confirmed the presence and location of erm(B) and tet(S), but genes providing tetracycline resistance in LbT16 were again not identified. In the genome of the multi-resistant strain LbE16, genes that might be involved in aminoglycoside (aadE, aphA-3, sat4) and virginiamycin [vat(E)] resistance were further found. The erm(B) gene but not tet(S) was transferred from Leuconostoc to Enterococcus faecalis both under laboratory conditions and in cheese. This study contributes to the characterization of AR in the Leuconostoc-Weissella group, provides evidence of the genetic basis of atypical resistances, and demonstrates the inter-species transfer of erythromycin resistance.     

Impact of Nisin-Activated Packaging on Microbiota of Beef Burgers during Storage

Beef burgers were stored at 4°C in a vacuum in nisin-activated antimicrobial packaging. Microbial ecology analyses were performed on samples collected between days 0 and 21 of storage to discover the population diversity. Two batches were analyzed using RNA-based denaturing gradient gel electrophoresis (DGGE) and pyrosequencing. The active packaging retarded the growth of the total viable bacteria and lactic acid bacteria. Culture-independent analysis by pyrosequencing of RNA extracted directly from meat showed that Photobacterium phosphoreum, Lactococcus piscium, Lactobacillus sakei, and Leuconostoc carnosum were the major operational taxonomic units (OTUs) shared between control and treated samples. Beta diversity analysis of the 16S rRNA sequence data and RNA-DGGE showed a clear separation between two batches based on the microbiota. Control samples from batch B showed a significant high abundance of some taxa sensitive to nisin, such as Kocuria rhizophila, Staphylococcus xylosus, Leuconostoc carnosum, and Carnobacterium divergens, compared to control samples from batch A. However, only from batch B was it possible to find a significant difference between controls and treated samples during storage due to the active packaging. Predicted metagenomes confirmed differences between the two batches and indicated that the use of nisin-based antimicrobial packaging can determine a reduction in the abundance of specific metabolic pathways related to spoilage. The present study aimed to assess the viable bacterial communities in beef burgers stored in nisin-based antimicrobial packaging, and it highlights the efficacy of this strategy to prolong beef burger shelf life.     

Substrate Preference in a Strain of Megasphaera elsdenii, a Ruminal Bacterium, and Its Implications in Propionate Production and Growth Competition

The NIAH 1102 strain of Megasphaera elsdenii utilized lactate in preference to glucose when the two substrates were present. Even when lactate was supplied to cells fermenting glucose, the cells switched substrate utilization from glucose to lactate and did not utilize glucose until lactate decreased to a low concentration (1 to 2 mM). Since substrate utilization was shifted gradually without intermittence, typical diauxic growth was not seen. The cyclic AMP content did not rise markedly with the shift in substrate utilization, suggesting that this nucleotide is not involved in the regulation of the shift. It was unlikely that propionate was produced from glucose, which was explicable by the fact that lactate racemase activity dropped rapidly with the exhaustion of lactate and cells actively fermenting glucose did not possess this enzyme. A coculture experiment indicated that M. elsdenii NIAH 1102 is overcome by Streptococcus bovis JB1 in the competition for glucose, mainly because M. elsdenii NIAH 1102 is obliged to utilize lactate produced by S. bovis JB1; i.e., glucose utilization by M. elsdenii NIAH 1102 is suppressed by the coexistence of S. bovis JB1.     

<Emphasis Type="Italic">Paenibacillus polymyxa</Emphasis> JB05-01-1 and its perspectives for food conservation and medical applications

The aim of this study was to isolate a novel bacterial strain with strong and broad spectrum antibacterial activity from a livestock feed prebiotic supplement. A novel strain, termed Paenibacillus polymyxa JB05-01-1, was isolated using traditional microbiological methods and identified on the basis of its phenotypic and biochemical properties as well as its 16S rRNA gene sequence. This strain was able to inhibit growth of gram-negative bacteria including Escherichia coli RR1, Pseudomonas fluorescens R73, Pantoea agglomerans BC1, Butyrivibrio fibrisolvens OR85, and Fibrobacter succinogenes. The above antagonism against the aforementioned bacteria was attributed to production of an antimicrobial substance(s) termed “JB05-01-1.” Its production was optimal during the stationary phase. JB05-01-1 has a molecular weight of 2.5 KDa, its mode of action is bactericidal, and the divalent cations, Ca²⁺ and Mn²⁺, reduced its lethality. The antibacterial activity was heat-stable and was effective at a pH range of 2–9. Enzymes like trypsin, α-chymotrypsin, and proteinase K have abolished the antibacterial activity of JB05-01-1 indicating a proteinaceous motif. This type of naturally occurring bacteria and inhibitory substance(s) could represent an additional value in livestock feed supplements. The natural presence of antibacterial activity indicates an opportunity to decrease the addition of antibiotics.     

Transmission of Pandora neoaphidis in the presence of co-occurring arthropods

Transmission of the entomopathogenic fungus Pandora neoaphidis to the nettle aphid Microlophium carnosum was assessed in the presence of arthropods that co-exist with the fungus within the habitat but do not compete for aphid hosts. The presence of a parasitoid significantly enhanced transmission, and transmission rates were similar for both enemy and non-enemy parasitoids. Although herbivory of nettle leaves by Peacock butterfly (Inchis io) caterpillars indirectly reduced the number of M. carnosum by >30% due to a reduction in leaf area for feeding, the addition of I. io significantly increased transmission of P. neoaphidis in the remaining aphids. It is likely that enhanced transmission in the presence of A. rhopalosiphii and I. io is due to disturbance and subsequent movement of the aphid, resulting in contact with conidia deposited on the leaf surface. The presence and impact of co-occurring arthropods should be taken into consideration when assessing the transmission of fungal entomopathogens.     

Antigenic Analysis of Virulent and Avirulent Strains of Trichomonas gallinae by Gel Diffusion Methods*

SYNOPSIS. Antigenic constitution of 5 Trichomonas gallinae strains and substrains was analyzed by gel diffusion technics. Fresh isolates of the very virulent JB and of an avirulent SG strain as well as avirulent substrains JBC and SGC, derived from JB and SG respectively by prolonged in vitro cultivation, were used in the experiments. An originally avirulent AG strain that was attenuated still further and lost its infectivity for pigeons during many years of serial transfers in nonliving media also was analyzed. Two major groups of antigens, A and B, were differentiated on the basis of precipitin line patterns formed in gel diffusion reactions involving the 5 strains and substrains and antisera prepared in rabbits against each of these trichomonad stocks. Group A was subdivided further into subgroups [A] and (A). JB, JBC, AG, and SGC trichomonads appeared to share all or nearly all antigens of both these subgroups, but AG strain contained some unique [A] and (A) antigens in addition to those which it had in common with the remaining 4 strains and substrains. Group B antigens were divided into 5 subgroups, B₁ to B₅. The complete B₁ antigenic complex was found in JB and JBC trichomonads and part of this complex was present also in SG strain and SGC substrain. In all instances, subgroup B₁ antigens stimulated production of specific antibodies in rabbits and combined with these antibodies present in immune sera. The complete B₂ antigenic complex was found only in JBC substrain. Some subgroup B₂ antigens were present also in JB trichomonads. Very few of these, however, were capable of stimulating antibody production in rabbits. The more numerous B₂ elements of JB strain that did not stimulate immunologic responses in rabbits, might be in the form of incomplete hapten-like antigens. All subgroup B₂ antigens found in JB strain represented only a portion of the B₂ complex associated with JBC substrain. Subgroup B₂ was characteristic of SG and SGC trichomonads, the latter substrain differing from the parental SG strain in the levels of both B₂ and B₁ antigens; these differences, however, were purely quantitative. JB strain reacted with some of subgroup B₃ antibodies present in SG and SGC antisera, but failed to stimulate antibody formation against any of these antigens in rabbits. The B₃ elements of JB trichomonads might be incomplete antigens. AG strain was characterized by having B₄ and B₅ antigenic complexes. The very small part of subgroup B₄, represented by a weak precipitin line in reactions between JB strain or JBC substrain and anti-AG serum, suggested the presence of some incomplete B₄ antigens in these trichomonads. Irrespective of whether freshly isolated avirulent strains or substrains attenuated by prolonged in vitro cultivation are examined by gel diffusion, such organisms are found richer in subgroup B antigens than the fully virulent JB trichomonads. All the results suggest that there may be a direct relationship between antigenic constitution and virulence of T. gallinae strains.     

Limonium failachicum (Plumbaginaceae) — New and so far the only endemic plant from Kuwait

Limonium failachicum, a new diploid (2n=18) species ofPlumbaginaceae, is described from Arabia. The species is taxonomically related toL. iranicum andL. carnosum. The new taxon is so far the only known endemic species for the State of Kuwait. Ecology and distribution of the new species is briefly discussed.     

Immunologic Analysis by Gel Diffusion Technics of the Effects of Prolonged Cultivation on Trichomonas gallinae*

SYNOPSIS. Antisera were developed in rabbits against 3 axenic lines of Trichomonas gallinae: JB(VI), the 6th isolate of the very virulent Jones' Barn strain, which was kept frozen in liquid nitrogen and had its full pathogenicity for pigeons when it was employed for immunization; JB(VI)C, a substrain derived from JB(VI), but attenuated during continuous in vitro cultivation for 1 year; and JB(V)C, a substrain of the 5th isolate of the Jones' Barn strain attenuated during continuous in vitro cultivation for over 3 years. All antisera were reacted on gel diffusion slides with varying concentrations of homologous and heterologous antigens. Two groups of precipitin bands, arbitrarily labeled A and B, were seen on the slides. Analysis of these bands revealed the common genetic makeup of the 3 trichomonad lines with respect to the group A bands. However, the group B antigenic system was strong in the attenuated JB(VI)C and JB(V)C substrains, and very weak in the fully pathogenic JB(VI) strain. These differences are discussed in the light of their possible relationship to pathogenicity.     

小麦赤霉病拮抗菌JB7的拮抗活性及鉴定

由禾谷镰刀菌(Fusarium graminearum, Fg)引起的赤霉病是限制小麦生产的主要病害之一。生物防治是一种高效且可持续的防治方法。【目的】从小麦种子内筛选具有抑制禾谷镰刀菌的菌株并对其生防潜力进行评估,为小麦赤霉病生防制剂的开发与利用提供菌种资源及理论支撑。【方法】采用平板对峙、孢子萌发法和无菌上清液抑菌试验筛选小麦种子内对禾谷镰刀菌具有拮抗活性的内生菌株;利用扫描电镜(scanning electron microscope, SEM)和共聚焦扫描电镜(confocal laser scanning microscope, CLSM)观察并分析无菌上清液对Fg的分生孢子形态、膜完整性以及胞内活性氧的影响;通过盆栽试验验证内生菌对小麦赤霉病的生防效果;应用二代Illumina HiSeq测序平台进行全基因组测序。【结果】从小麦种子中分离出一株高效抑制Fg生长的内生菌株JB7,其衰亡期无菌上清液对Fg孢子萌发抑制率高达85.23%。菌株JB7的无菌上清液使Fg孢子表面凹陷,破坏其细胞膜,造成核酸和蛋白质的渗漏,诱导Fg菌丝活性氧的累积,引起Fg菌丝可溶性蛋白和丙二醛含量的显著升高。该菌株具有分泌蛋白酶、纤维素酶、葡聚糖酶和产铁载体的能力。盆栽试验表明菌株JB7能显著降低小麦赤霉病的病情指数(P<0.05)。经全基因组学鉴定为甲基营养型芽孢杆菌(Bacillus methylotrophicus) JB7,该菌株基因组中含有12个抑菌功能的次级代谢产物合成基因簇。【结论】菌株JB7能抑制禾谷镰刀菌的生长,对小麦赤霉病有较强的防效,可作为生物防治小麦赤霉病的候选菌株。     

Potential value of the fibre nettle <Emphasis Type="Italic">Urtica dioica</Emphasis> as a resource for the nettle aphid <Emphasis Type="Italic">Microlophium carnosum</Emphasis> and its insect and fungal natural enemies

The perennial stinging nettle (Urtica dioica L.) is a wild plant that provides resources for aphid natural enemies and can therefore benefit crop protection. Stinging nettles producing approximately three times more fibre than Standard nettles are under commercial development for fibre production. Here we assess the relative value of Austrian Clone 2, a high fibre nettle variety, as a resource for the nettle aphid Microlophium carnosum (Buckton) and its associated natural enemies. The intrinsic rate of increase of M. carnosum cultured on Clone 2 was not different to that on a Standard nettle nor was there an effect of nettle variety on the susceptibility of this aphid to the entomopathogenic fungus Pandora neoaphidis (Remaudière and Hennebert) Humber. The development time of the aphidophagous predator Chrysoperla carnea (Stephens) was not affected by the nettle variety on which M. carnosum was cultured whilst the parasitoid Aphidius ervi (Haliday) fed on honeydew produced by M. carnosum infesting both varieties of nettle. Fibre nettle Clone 2 is therefore able to support non-pest aphids and their natural enemies and, if grown widely in the future, may be useful within conservation biological control and a tool within integrated pest management.     

Photobacterium carnosum sp. nov., isolated from spoiled modified atmosphere packaged poultry meat

Analysis of spoilage-associated microbiota of modified-atmosphere packaged poultry meat revealed four different bacterial isolates that could not be assigned to known species. They showed a Gram-negative staining behavior, were facultatively aerobic, non-motile with variable cell morphology. Phylogenetic analysis of 16S rDNA and gyrB, rpoD and recA revealed a distinct lineage within the genus Photobacterium with Photobacterium (P.) iliopiscarium DSM 9896^T, P. phosphoreum DSM 15556^T, P. kishitanii DSM 19954^T, P. piscicola LMG 27681^T and P. aquimaris DSM 23343^T as closest relatives.The designated type strain TMW 2.2021^T is non-luminous and grew at 0–20 °C (optimum 10–15 °C), within pH 5.0–8.5 (optimum 6–8) and in the presence of 0.5–3% (w/v) NaCl (optimum 1%). Major cellular fatty acids of TMW 2.2021^T were summed feature 3 (C_16:1ω7c/iso-C₁₅ 3-OH), C_16:0, C_18:1ω7c and summed feature 2 (C_12:0 aldehyde and C_10.928 unknown). Quinone analysis revealed Q-8 as sole respiratory ubiquinone. The genome of TMW 2.2021^T has a size of 4.56 Mb and a G + C content of 38.49 mol%. The ANI value between TMW 2.2021^T and the type strain of closest relative P. iliopiscarium DSM 9896^T was 91.43%. Fingerprinting on the base of M13-RAPD-PCR band pattern and MALDI-TOF MS profiles allowed intraspecies differentiation between our isolates but also supported their distinct lineage to a novel species. Based on phylogenetic, genomic, phenotypic and chemotaxonomic data, strain TMW 2.2021^T and further strains represent a novel species of the genus Photobacterium, for which the name Photobacterium carnosum sp. nov. is proposed. The type strain is TMW 2.2021^T (=DSM 105454^T = CECT 9394^T).     

Jolkinolide B induces cell cycle arrest and apoptosis in MKN45 gastric cancer cells and inhibits xenograft tumor growth in vivo

Gastric cancer is one of the most common digestive carcinomas throughout the world and represents high mortality. There is an urgent quest for seeking a novel and efficient antigastric cancer drug. Euphorbia fischeriana Steud had long been used as a traditional Chinese medicine for the treatment of cancer. According to the basic theory of traditional Chinese medicine, its antitumor mechanism is ‘to combat poison with poison’. However, its effective material foundation of it is still ambiguous. In our previous work, we studied the chemical constituents of E. fischeriana Steud. Jolkinolide B (JB) is an ent-abietane-type diterpenoid we isolated from it. The purpose of the present study was to investigate the antigastric effect and mechanism of JB. Results revealed that JB could suppress the proliferation of MKN45 cells in vitro and inhibit MKN45 xenograft tumor growth in nude mice in vivo. We further investigated its anticancer mechanism. On the one hand, JB caused DNA damage in gastric cancer MKN45 cells and induced the S cycle arrest by activating the ATR-CHK1-CDC25A-Cdk2 signaling pathway, On the other hand, JB induced MKN45 cells apoptosis through the mitochondrial pathway, and ultimately effectively inhibited the growth of gastric cancer cells. These results suggest that JB appears to be a promising candidate drug with antigastric cancer activity and warrants further research.