Stress tolerant virulent strains of Cronobacter sakazakii from food

Background

Cronobacter sakazakii is considered as an emerging foodborne pathogen. The aim of this study was to isolate and characterize virulent strains of Cronobacter sakazakii from food samples of Bangladesh.

Result

Six (6) Cronobacter sakazakii was isolated and identified from 54 food samples on the basis of biochemical characteristics, sugar fermentation, SDS-PAGE of whole cell protein, plasmid profile and PCR of Cronobacter spp. specific genes (esak, gluA, zpx, ompA, ERIC, BOX-AIR) and sequencing. These strains were found to have moderately high antibiotic resistance against common antibiotics and some are ESBL producer. Most of the C. sakazakii isolates were capable of producing biofilm (strong biofilm producer), extracellular protease and siderophores, curli expression, haemolysin, haemagglutinin, mannose resistant haemagglutinin, had high cell surface hydrophobicity, significant resistance to human serum, can tolerate high concentration of salt, bile and DNase production. Most of them produced enterotoxins of different molecular weight. The isolates pose significant serological cross-reactivity with other gram negative pathogens such as serotypes of Salmonella spp., Shigella boydii, Shigella sonnei, Shigella flexneri and Vibrio cholerae. They had significant tolerance to high temperature, low pH, dryness and osmotic stress.

Conclusion

Special attention should be given in ensuring hygiene in production and post-processing to prevent contamination of food with such stress-tolerant virulent Cronobacter sakazakii.

Electronic supplementary material

The online version of this article (doi:10.1186/0717-6287-47-63) contains supplementary material, which is available to authorized users.     

Use of rpoB gene sequence analysis for phylogenetic identification of Cronobacter species

Here, we report the RNA polymerase beta-subunit gene (rpoB) as a new molecular marker for the identification of the Cronobacter species. The results indicated that members of the Cronobacter genus are more easily discriminated by rpoB sequencing than 16S rRNA sequencing, and reliable identification could be achieved by rpoB gene sequence comparison.     

Complete genome sequence of Streptococcus agalactiae strain SA20-06, a fish pathogen associated to meningoencephalitis outbreaks

Streptococcus agalactiae (Lancefield group B; GBS) is the causative agent of meningoencephalitis in fish, mastitis in cows, and neonatal sepsis in humans. Meningoencephalitis is a major health problem for tilapia farming and is responsible for high economic losses worldwide. Despite its importance, the genomic characteristics and the main molecular mechanisms involved in virulence of S. agalactiae isolated from fish are still poorly understood. Here, we present the genomic features of the 1,820,886 bp long complete genome sequence of S. agalactiae SA20-06 isolated from a meningoencephalitis outbreak in Nile tilapia (Oreochromis niloticus) from Brazil, and its annotation, consisting of 1,710 protein-coding genes (excluding pseudogenes), 7 rRNA operons, 79 tRNA genes and 62 pseudogenes.     

Non-contiguous finished genome sequence of the opportunistic oral pathogen Prevotella multisaccharivorax type strain (PPPA20)

Prevotella multisaccharivorax Sakamoto et al. 2005 is a species of the large genus Prevotella, which belongs to the family Prevotellaceae. The species is of medical interest because its members are able to cause diseases in the human oral cavity such as periodontitis, root caries and others. Although 77 Prevotella genomes have already been sequenced or are targeted for sequencing, this is only the second completed genome sequence of a type strain of a species within the genus Prevotella to be published. The 3,388,644 bp long genome is assembled in three non-contiguous contigs, harbors 2,876 protein-coding and 75 RNA genes and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.     

Non contiguous-finished genome sequence and description of Enterobacter massiliensis sp. nov.

Enterobacter massiliensis strain JC163^T sp. nov. is the type strain of E. massiliensis sp. nov., a new species within the genus Enterobacter. This strain, whose genome is described here, was isolated from the fecal flora of a healthy Senegalese patient. E. massiliensis is an aerobic rod. Here we describe the features of this organism, together with the complete genome sequence and annotation. The 4,922,247 bp long genome (1 chromosome but no plasmid) exhibits a G+C content of 55.1% and contains 4,644 protein-coding and 80 RNA genes, including 5 rRNA genes.     

A new cost-effective, selective and differential medium for the isolation of Cronobacter spp

The objective of the present study was to develop a new selective, differential and cost-effective medium (Kim and Rhee — KR-medium) for the isolation of Cronobacter spp. In this new medium, which contained salicin as a differential agent, Cronobacter spp. generated typical colonies with characteristic violet-colored centers surrounded by a transparent to opalescent border, and the growth of other microorganisms (40 strains) was inhibited or produced visually distinguishable colonies. Using healthy and heat- and desiccation-injured cells, the quantity of nutrients was adjusted to determine the optimal recovery rate, selectivity, differentiation and cost-effectiveness. Peptone and salicin concentrations were established as 10 and 8 g/L, respectively. The KR medium was then validated using salicin fermenting organisms, including Cronobacter spp. (52 strains), Enterobacter cloacae (50 strains) and Klebsiella pneumonia (10 strains) isolated from clinical and food specimens. All strains of Cronobacter spp. produced typical colonies and other salicin fermenting organisms were easily distinguishable from Cronobacter spp. with the exception of 2 E. cloacae strains. The verification of KR medium was carried out in powdered infant formula artificially inoculated with healthy, heat-injured, and desiccation-injured Cronobacter spp. and the expected typical colonies were appeared. The KR medium had a high specificity (98%) and sensitivity (100%), with no false-negative results. Moreover, we show that the cost of the KR medium is much lower than that of other selective and differential media. The use of the KR medium for the selective isolation of Cronobacter spp. in laboratories and food industry settings may therefore lessen the financial burden of Cronobacter spp. detection.     

Complete genome sequence of Arcanobacterium haemolyticum type strain (11018)

Arcanobacterium haemolyticum (ex MacLean et al. 1946) Collins et al. 1983 is the type species of the genus Arcanobacterium, which belongs to the family Actinomycetaceae. The strain is of interest because it is an obligate parasite of the pharynx of humans and farm animal; occasionally, it causes pharyngeal or skin lesions. It is a Gram-positive, nonmotile and non-sporulating bacterium. The strain described in this study was isolated from infections amongst American soldiers of certain islands of the North and West Pacific. This is the first completed sequence of a member of the genus Arcanobacterium and the ninth type strain genome from the family Actinomycetaceae. The 1,986,154 bp long genome with its 1,821 protein-coding and 64 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.     

Complete genome sequence of Methanoplanus petrolearius type strain (SEBR 4847)

Segniliparus rotundus Butler 2005 is the type species of the genus Segniliparus, which is currently the only genus in the corynebacterial family Segniliparaceae. This family is of large interest because of a novel late-emerging genus-specific mycolate pattern. The type strain has been isolated from human sputum and is probably an opportunistic pathogen. Here we describe the features of this organism, together with the complete genome sequence and annotation. This is the first completed genome sequence of the family Segniliparaceae. The 3,157,527 bp long genome with its 3,081 protein-coding and 52 RNA genes is part of the Genomic Encyclopedia of Bacteria and Archaea project.     

The complete genome sequence of Clostridium indolis DSM 755T

Clostridium indolis DSM 755^T is a bacterium commonly found in soils and the feces of birds and mammals. Despite its prevalence, little is known about the ecology or physiology of this species. However, close relatives, C. saccharolyticum and C. hathewayi, have demonstrated interesting metabolic potentials related to plant degradation and human health. The genome of C. indolis DSM 755^T reveals an abundance of genes in functional groups associated with the transport and utilization of carbohydrates, as well as citrate, lactate, and aromatics. Ecologically relevant gene clusters related to nitrogen fixation and a unique type of bacterial microcompartment, the CoAT BMC, are also detected. Our genome analysis suggests hypotheses to be tested in future culture based work to better understand the physiology of this poorly described species.     

Complete genome sequence of Riemerella anatipestifer type strain (ATCC 11845)

Riemerella anatipestifer (Hendrickson and Hilbert 1932) Segers et al. 1993 is the type species of the genus Riemerella, which belongs to the family Flavobacteriaceae. The species is of interest because of the position of the genus in the phylogenetic tree and because of its role as a pathogen of commercially important avian species worldwide. This is the first completed genome sequence of a member of the genus Riemerella. The 2,155,121 bp long genome with its 2,001 protein-coding and 51 RNA genes consists of one circular chromosome and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.     

Complete genome sequence of Odoribacter splanchnicus type strain (1651/6)

Odoribacter splanchnicus (Werner et al. 1975) Hardham et al. 2008 is the type species of the genus Odoribacter, which belongs to the family Porphyromonadaceae in the order 'Bacteroidales'. The species is of interest because members of the Odoribacter form an isolated cluster within the Porphyromonadaceae. This is the first completed genome sequence of a member of the genus Odoribacter and the fourth sequence from the family Porphyromonadaceae. The 4,392,288 bp long genome with its 3,672 protein-coding and 74 RNA genes and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.     

Complete genome sequence of Enterobacter sp. IIT-BT 08: A potential microbial strain for high rate hydrogen production

Enterobacter sp. IIT-BT 08 belongs to Phylum: Proteobacteria, Class: Gammaproteobacteria, Order: Enterobacteriales, Family: Enterobacteriaceae. The organism was isolated from the leaves of a local plant near the Kharagpur railway station, Kharagpur, West Bengal, India. It has been extensively studied for fermentative hydrogen production because of its high hydrogen yield. For further enhancement of hydrogen production by strain development, complete genome sequence analysis was carried out. Sequence analysis revealed that the genome was linear, 4.67 Mbp long and had a GC content of 56.01%. The genome properties encode 4,393 protein-coding and 179 RNA genes. Additionally, a putative pathway of hydrogen production was suggested based on the presence of formate hydrogen lyase complex and other related genes identified in the genome. Thus, in the present study we describe the specific properties of the organism and the generation, annotation and analysis of its genome sequence as well as discuss the putative pathway of hydrogen production by this organism.     

Typing of Campylobacter jejuni and Campylobacter coli isolated from live broilers and retail broiler meat by flaA-RFLP, MLST, PFGE and REP-PCR

We analyzed 100 Campylobacter spp. isolates (C. jejuni and C. coli) from Grenada, Puerto Rico and Alabama, which were collected from live broilers or retail broiler meat. We analyzed these isolates with four molecular typing methods: restriction fragment length polymorphism of the flaA gene (flaA-RFLP), multilocus sequence typing (MLST), pulsed-field gel electrophoresis (PFGE), and automated repetitive extragenic palindromic polymerase chain reaction (REP-PCR) using the DiversiLab system. All methods performed similarly for the typing of C. jejuni and C. coli. The DNA extraction method appears to influence the results obtained with REP-PCR. This method was better for the typing of C. jejuni than C. coli, however both REP-PCR and flaA-RFLP generated types that were indistinguishable between C. jejuni and C. coli and appeared to be random, without any relationship to species, location, or source of isolates. PFGE and MLST generated typing results that had a better correlation with the geographic location of the isolates and showed higher concordance with the Wallace coefficient. The adjusted Rand coefficient did not show higher concordance among the methods, although the PFGE/MLST combination exhibited the highest concordance. PFGE and MLST revealed a better discriminatory power for C. coli isolates than REP-PCR or flaA-RFLP. The use of readily available online tools to calculate the confidence interval of the Simpson's index of diversity and the adjusted Rand and Wallace coefficients helped estimate the discriminatory power of typing methods. Further studies using different C. jejuni and C. coli strains may expand our understanding of the benefits and limitations of each of these typing methods for epidemiological studies of Campylobacter spp.     

Complete genome sequence of Tsukamurella paurometabola type strain (no. 33)

Tsukamurella paurometabola corrig. (Steinhaus 1941) Collins et al. 1988 is the type species of the genus Tsukamurella, which is the type genus to the family Tsukamurellaceae. The species is not only of interest because of its isolated phylogenetic location, but also because it is a human opportunistic pathogen with some strains of the species reported to cause lung infection, lethal meningitis, and necrotizing tenosynovitis. This is the first completed genome sequence of a member of the genus Tsukamurella and the first genome sequence of a member of the family Tsukamurellaceae. The 4,479,724 bp long genome contains a 99,806 bp long plasmid and a total of 4,335 protein-coding and 56 RNA genes, and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.     

Complete genome sequence of Nocardiopsis dassonvillei type strain (IMRU 509)

Nocardiopsis dassonvillei (Brocq-Rousseau 1904) Meyer 1976 is the type species of the genus Nocardiopsis, which in turn is the type genus of the family Nocardiopsaceae. This species is of interest because of its ecological versatility. Members of N. dassonvillei have been isolated from a large variety of natural habitats such as soil and marine sediments, from different plant and animal materials as well as from human patients. Moreover, representatives of the genus Nocardiopsis participate actively in biopolymer degradation. This is the first complete genome sequence in the family Nocardiopsaceae. Here we describe the features of this organism, together with the complete genome sequence and annotation. The 6,543,312 bp long genome consist of a 5.77 Mbp chromosome and a 0.78 Mbp plasmid and with its 5,570 protein-coding and 77 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.     

Complete genome sequence of Bacteroides helcogenes type strain (P 36-108)

Bacteroides helcogenes Benno et al. 1983 is of interest because of its isolated phylogenetic location and, although it has been found in pig feces and is known to be pathogenic for pigs, occurrence of this bacterium is rare and it does not cause significant damage in intensive animal husbandry. The genome of B. helcogenes P 36-108(T) is already the fifth completed and published type strain genome from the genus Bacteroides in the family Bacteroidaceae. The 3,998,906 bp long genome with its 3,353 protein-coding and 83 RNA genes consists of one circular chromosome and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.     

Host and pathogen ecology drive the seasonal dynamics of a fungal disease,white-nose syndrome

Seasonal patterns in pathogen transmission can influence the impact of disease on populations and the speed of spatial spread. Increases in host contact rates or births drive seasonal epidemics in some systems, but other factors may occasionally override these influences. White-nose syndrome, caused by the emerging fungal pathogen Pseudogymnoascus destructans, is spreading across North America and threatens several bat species with extinction. We examined patterns and drivers of seasonal transmission of P. destructans by measuring infection prevalence and pathogen loads in six bat species at 30 sites across the eastern United States. Bats became transiently infected in autumn, and transmission spiked in early winter when bats began hibernating. Nearly all bats in six species became infected by late winter when infection intensity peaked. In summer, despite high contact rates and a birth pulse, most bats cleared infections and prevalence dropped to zero. These data suggest the dominant driver of seasonal transmission dynamics was a change in host physiology, specifically hibernation. Our study is the first, to the best of our knowledge, to describe the seasonality of transmission in this emerging wildlife disease. The timing of infection and fungal growth resulted in maximal population impacts, but only moderate rates of spatial spread.     

Structural and sequence diversity of the transposon Galileo in the Drosophila willistoni genome

Background

Galileo is one of three members of the P superfamily of DNA transposons. It was originally discovered in Drosophila buzzatii, in which three segregating chromosomal inversions were shown to have been generated by ectopic recombination between Galileo copies. Subsequently, Galileo was identified in six of 12 sequenced Drosophila genomes, indicating its widespread distribution within this genus. Galileo is strikingly abundant in Drosophila willistoni, a neotropical species that is highly polymorphic for chromosomal inversions, suggesting a role for this transposon in the evolution of its genome.

Results

We carried out a detailed characterization of all Galileo copies present in the D. willistoni genome. A total of 191 copies, including 133 with two terminal inverted repeats (TIRs), were classified according to structure in six groups. The TIRs exhibited remarkable variation in their length and structure compared to the most complete copy. Three copies showed extended TIRs due to internal tandem repeats, the insertion of other transposable elements (TEs), or the incorporation of non-TIR sequences into the TIRs. Phylogenetic analyses of the transposase (TPase)-encoding and TIR segments yielded two divergent clades, which we termed Galileo subfamilies V and W. Target-site duplications (TSDs) in D. willistoni Galileo copies were 7- or 8-bp in length, with the consensus sequence GTATTAC. Analysis of the region around the TSDs revealed a target site motif (TSM) with a 15-bp palindrome that may give rise to a stem-loop secondary structure.

Conclusions

There is a remarkable abundance and diversity of Galileo copies in the D. willistoni genome, although no functional copies were found. The TIRs in particular have a dynamic structure and extend in different ways, but their ends (required for transposition) are more conserved than the rest of the element. The D. willistoni genome harbors two Galileo subfamilies (V and W) that diverged ~9 million years ago and may have descended from an ancestral element in the genome. Galileo shows a significant insertion preference for a 15-bp palindromic TSM.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-792) contains supplementary material, which is available to authorized users.     

Comprehensive molecular,genomic and phenotypic analysis of a major clone of Enterococcus faecalis MLST ST40

Background

Enterococcus faecalis is a multifaceted microorganism known to act as a beneficial intestinal commensal bacterium. It is also a dreaded nosocomial pathogen causing life-threatening infections in hospitalised patients. Isolates of a distinct MLST type ST40 represent the most frequent strain type of this species, distributed worldwide and originating from various sources (animal, human, environmental) and different conditions (colonisation/infection). Since enterococci are known to be highly recombinogenic we determined to analyse the microevolution and niche adaptation of this highly distributed clonal type.

Results

We compared a set of 42 ST40 isolates by assessing key molecular determinants, performing whole genome sequencing (WGS) and a number of phenotypic assays including resistance profiling, formation of biofilm and utilisation of carbon sources. We generated the first circular closed reference genome of an E. faecalis isolate D32 of animal origin and compared it with the genomes of other reference strains. D32 was used as a template for detailed WGS comparisons of high-quality draft genomes of 14 ST40 isolates. Genomic and phylogenetic analyses suggest a high level of similarity regarding the core genome, also demonstrated by similar carbon utilisation patterns. Distribution of known and putative virulence-associated genes did not differentiate between ST40 strains from a commensal and clinical background or an animal or human source. Further analyses of mobile genetic elements (MGE) revealed genomic diversity owed to: (1) a modularly structured pathogenicity island; (2) a site-specifically integrated and previously unknown genomic island of 138 kb in two strains putatively involved in exopolysaccharide synthesis; and (3) isolate-specific plasmid and phage patterns. Moreover, we used different cell-biological and animal experiments to compare the isolate D32 with a closely related ST40 endocarditis isolate whose draft genome sequence was also generated. D32 generally showed a greater capacity of adherence to human cell lines and an increased pathogenic potential in various animal models in combination with an even faster growth in vivo (not in vitro).

Conclusion

Molecular, genomic and phenotypic analysis of representative isolates of a major clone of E. faecalis MLST ST40 revealed new insights into the microbiology of a commensal bacterium which can turn into a conditional pathogen.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1367-x) contains supplementary material, which is available to authorized users.