Int J Med Microbiol [journal]
- Stability of the cargo regions of the cfr-carrying, multiresistance plasmid pSP01 from Staphylococcus epidermidis. [JOURNAL ARTICLE]
- Int J Med Microbiol 2016 Aug 17.
This study investigated the stability or instability - i.e. the ability or inability to undergo excision in circular form - of the four cargo regions (cr1 to cr4) of the novel cfr-carrying, multiresistance plasmid pSP01, arboured by a clinical Staphylococcus epidermidis isolate. Only cr4 proved unstable. The stability of cr1 and cr2 was substantially expected. Insertion sequences (ISs) played an important role in the stability of cr3 (the cfr gene context) and in the instability of cr4. Whereas the stability of cfr genetic contexts is associated with the presence of a single IS copy (istAS-istBS in cr3), their instability is associated with two identical, flanking ISs with the same orientation. cr4 is bracketed between two identical IS257 elements, and appears to behave as a composite transposon. Its instability is of interest because of the existence of a closely related cfr plasmid from S. epidermidis (pSP01.1) that differs from pSP01 only by the lack of cr4. An integration/recombination mechanism is suggested to explain how cr4 may have moved to pSP01.1 to form pSP01.
- Genetic features of livestock-associated Staphylococcus aureus ST9 isolates from Chinese pigs that carry the lsa(E) gene for quinupristin/dalfopristin resistance. [JOURNAL ARTICLE]
- Int J Med Microbiol 2016 Aug 10.
Whole-genome sequencing (WGS) was used to investigate the genetic features of the recently identified lsa(E) gene in porcine S. aureus ST9 isolates. Three quinupristin/dalfopristin-resistant isolates harboring the lsa(E) gene (two MRSA and one MSSA) were sequenced. Phylogenetic analysis of 184S. aureus genomes showed that ST9 porcine isolates belong to a distinct sequence cluster. Further analysis showed that all isolates were deficient in the recently described type IV restriction-modification system and SCCmec type XII was identified in the two MRSA isolates, which included a rare class C2 mec gene complex. A 24kb ΨSCC fragment was found in the MRSA and MSSA isolates sharing 99% nucleotide sequence homology with the ΨSCCJCSC6690 (O-2) element of a ST9 MRSA isolate from Thailand (accession number AB705453). Comparison of these ST9 isolates with 181 publically available S. aureus genomes identified 24 genes present in all (100%) ST9 isolates, that were absent from the most closely related human isolate. Our analysis suggests that the sequenced quinupristin/dalfopristin-resistant ST9 lineage represent a reservoir of mobile genetic elements associated with resistance and virulence features.
- Molecular characterisation of Czech Clostridium difficile isolates collected in 2013-2015. [JOURNAL ARTICLE]
- Int J Med Microbiol 2016 Aug 5.
Clostridium difficile is a leading nosocomial pathogen and molecular typing is a crucial part of monitoring its occurrence and spread. Over a three-year period (2013-2015), clinical C. difficile isolates from 32 Czech hospitals were collected for molecular characterisation. Of 2201 C. difficile isolates, 177 (8%) were non-toxigenic, 2024 (92%) were toxigenic (tcdA and tcdB) and of these, 677 (33.5%) carried genes for binary toxin production (cdtA, cdtB). Capillary-electrophoresis (CE) ribotyping of the 2201 isolates yielded 166 different CE-ribotyping profiles, of which 53 were represented by at least two isolates for each profile. Of these, 29 CE-ribotyping patterns were common to the Leeds-Leiden C. difficile reference strain library and the WEBRIBO database (83.7% isolates), and 24 patterns were recognized only by the WEBRIBO database (11.2% isolates). Isolates belonging to these 53 CE-ribotyping profiles comprised 94.9% of all isolates. The ten most frequent CE-ribotyping profiles were 176 (n=588, 26.7%), 001 (n=456, 20.7%), 014 (n=176, 8%), 012 (n=127, 5.8%), 017 (n=85, 3.9%), 020 (n=68, 3.1%), 596 (n=55, 2.5%), 002-like (n=45, 2.1%), 010 (n=35, 1.6%) and 078 (n=34, 1.6%). Multi-locus sequence typing (MLST) of seven housekeeping genes performed in one isolate of each of 53 different CE-ribotyping profiles revealed 40 different sequence types (STs). We conclude that molecular characterisation of Czech C. difficile isolates revealed a high diversity of CE-ribotyping profiles; the prevailing RTs were 001 (20.7%) and 176 (027-like, 26.7%).
- Colicins U and Y inhibit growth of Escherichia coli strains via recognition of conserved OmpA extracellular loop 1. [JOURNAL ARTICLE]
- Int J Med Microbiol 2016 Aug 2.
Interactions of colicins U and Y with the OmpA (Outer membrane protein A) receptor molecule were studied using site-directed mutagenesis and colicin binding assay. A systematic mutagenesis of the colicin-susceptible OmpA sequence from Escherichia coli (OmpAEC) to the colicin-resistant OmpA sequence from Serratia marcescens (OmpASM) was performed in regions corresponding to extracellular OmpA loops 1-4. Susceptibility to colicins U and Y was significantly affected by the OmpA mutation in loop 1. As with functional analysis, a decrease in binding capacity of His-tagged colicin U was found for recombinant OmpA with a mutated segment in loop 1 compared to control OmpAEC. To verify the importance of the identified amino acid residues in OmpA loop 1, we introduced loop 1 from OmpAEC into OmpASM, which resulted in the substantial increase of susceptibility to colicins U and Y. In addition, colicins U and Y were tested against a panel of 118 bacteriocin non-producing strains of four Escherichia species, including E. coli (39 strains), E. fergusonii (10 strains), E. hermannii (42 strains), and E. vulneris (27 strains). A majority (82%) of E. coli strains was susceptible to colicins U and Y. Interestingly, colicins U and Y also inhibited all of the 30 tested multidrug-resistant E. coli O25b-ST131 isolates. These findings, together with the fact that OmpA loop 1 is important for bacterial virulence and is evolutionary conserved, offer the potential of using colicins U and Y as specific anti-OmpA loop 1 directed antibacterial proteins.
- A hypervariable genomic island identified in clinical and environmental Mycobacterium avium subsp. hominissuis isolates from Germany. [JOURNAL ARTICLE]
- Int J Med Microbiol 2016 Jul 18.
Mycobacterium avium subsp. hominissuis (MAH) is an opportunistic human pathogen widespread in the environment. Genomic islands (GI)s represent a part of the accessory genome of bacteria and influence virulence, drug-resistance or fitness and trigger bacterial evolution. We previously identified a novel GI in four MAH genomes. Here, we further explored this GI in a larger collection of MAH isolates from Germany (n=41), including 20 clinical and 21 environmental isolates. Based on comparative whole genome analysis, we detected this GI in 39/41 (95.1%) isolates. Although all these GIs integrated in the same insertion hotspot, there is high variability in the genetic structure of this GI: eight different types of GI have been identified, designated A-H (sized 6.2-73.3kb). These GIs were arranged as single GI (23/41, 56.1%), combination of two different GIs (14/41, 34.1%) or combination of three different GIs (2/41, 4.9%) in the insertion hotspot. Moreover, two GI types shared more than 80% sequence identity with sequences of M. canettii, responsible for Tuberculosis. A total of 253 different genes were identified in all GIs, among which the previously documented virulence-related genes mmpL10 and mce. The diversity of the GI and the sequence similarity with other mycobacteria suggests cross-species transfer, involving also highly pathogenic species. Shuffling of potential virulence genes such as mmpL10 via this GI may create new pathogens that can cause future outbreaks.
- Heterologous Pseudomonas aeruginosa O-antigen delivery using a Salmonella enterica serovar Typhimurium wecA mutant strain. [JOURNAL ARTICLE]
- Int J Med Microbiol 2016 Jun 28.
There is a broad interest in adapting live vaccine strains (LVS) for use as recombinant vaccines that can deliver heterologous antigens. The Salmonella enterica serovar Typhimurium SL1344 ΔwecA LVS contains a mutation in wecA that abrogates production of Enterobacterial common antigen. This ΔwecA strain is attenuated in vivo, persistently colonizes the host, and protects against both wild type and cross-Salmonella serovar lethal challenge in a murine model of salmonellosis. Given these characteristics, we hypothesized that the SL1344 ΔwecA strain could be used as a carrier for heterologous antigen expression. To test this hypothesis, SL1344 ΔwecA was engineered to express the Pseudomonas aeruginosa O11 O-antigen gene cluster. Intraperitoneal (IP) but not oral immunization of BALB/c mice with the heterologous expression strain protected against lethal P. aeruginosa intranasal (IN) challenge. Furthermore, IP immunization resulted in P. aeruginosa O11-specific Ig and IgG antibody production. Functional analysis of sera collected from the IP immunized mice showed antibody-mediated agglutination and opsonophagocytic activity against P. aeruginosa. En masse, these results indicate that the S. Typhimurium SL1344 ΔwecA strain expressing the P. aeruginosa O11 O-antigen gene cluster is able to induce a humoral immune response and to protect against lethal P. aeruginosa challenge. As such, the S. Typhimurium SL1344 ΔwecA LVS can likely serve as a vehicle for expression of a wide variety of heterologous antigens as a means to create recombinant vaccines.
- Nutritional influences of overfeeding on experimental outcomes in laboratory mice: consequences for gut microbiota and other functional studies. [Journal Article]
- Int J Med Microbiol 2016 Aug; 306(5):328-33.
Data from literature suggests that laboratory mice are often overfed and malnourished. This might have several reasons, including: (i) we usually offer an ad libitum diet, which is not the natural way of feeding for a wild mouse; (ii) many commercial diets we use contain rather high amounts of carbohydrates, particularly of sugars, and low amounts of fat; and (iii) laboratory mice live in a warm and constricted environment in which energy expenditure is lower than in the wild. Such selective or global overfeeding in laboratory mice, which resembles the widespread overfeeding in humans, although it does not always result in overweight, likely affects a number of outcome variables analyzed in laboratory mice, such as microbiota composition and function, metabolic alterations, longevity, intestinal permeability and inflammation. Therefore, a careful selection of experimental diets and their way of administration, as well as detailed documentation, is mandatory in order to understand and compare scientific data obtained from different mouse experiments.
- Propionibacterium acnes inhibits FOXM1 and induces cell cycle alterations in human primary prostate cells. [JOURNAL ARTICLE]
- Int J Med Microbiol 2016 Jun 28.
Propionibacterium acnes has been detected in diseased human prostate tissue, and cell culture experiments suggest that the bacterium can establish a low-grade inflammation. Here, we investigated its impact on human primary prostate epithelial cells. Microarray analysis confirmed the inflammation-inducing capability of P. acnes but also showed deregulation of genes involved in the cell cycle. qPCR experiments showed that viable P. acnes downregulates a master regulator of cell cycle progression, FOXM1. Flow cytometry experiments revealed that P. acnes increases the number of cells in S-phase. We tested the hypothesis that a P. acnes-produced berninamycin-like thiopeptide is responsible for this effect, since it is related to the FOXM1 inhibitor siomycin. The thiopeptide biosynthesis gene cluster was strongly expressed; it is present in subtype IB of P. acnes, but absent from type IA, which is most abundant on human skin. A knock-out mutant lacking the gene encoding the berninamycin-like peptide precursor was unable to downregulate FOXM1 and to halt the cell cycle. Our study reveals a novel host cell-interacting activity of P. acnes.
- Differential compartmentalization of Streptococcus pyogenes virulence factors and host protein binding properties as a mechanism for host adaptation. [JOURNAL ARTICLE]
- Int J Med Microbiol 2016 Jun 29.
Streptococcus pyogenes is an important human pathogen responsible for substantial morbidity and mortality worldwide. Although S. pyogenes is a strictly human pathogen with no other known animal reservoir, several murine infection models exist to explore different aspects of the bacterial pathogenesis. Inoculating mice with wild-type S. pyogenes strains can result in the generation of new bacterial phenotypes that are hypervirulent compared to the original inoculum. In this study, we used a serial mass spectrometry based proteomics strategy to investigate if these hypervirulent strains have an altered distribution of virulence proteins across the intracellular, surface associated and secreted bacterial compartments and if any change in compartmentalization can alter the protein-protein interaction network between bacteria and host proteins. Quantitative analysis of the S. pyogenes surface and secreted proteomes revealed that animal passaged strains are associated with significantly higher amount of virulence factors on the bacterial surface and in the media. This altered virulence factor compartmentalization results in increased binding of several mouse plasma proteins to the bacterial surface, a trend that was consistent for mouse plasma from several different mouse strains. In general, both the wild-type strain and animal passaged strain were capable of binding high amounts of human plasma proteins. However, compared to the non-passaged strains, the animal passaged strains displayed an increased ability to bind mouse plasma proteins, in particular for M protein binders, indicating that the increased affinity for mouse blood plasma proteins is a consequence of host adaptation of this pathogen to a new host. In conclusion, plotting the total amount of virulence factors against the total amount of plasma proteins associated to the bacterial surface could clearly separate out animal passaged strains from wild type strains indicating a virulence model that could predict the virulence of a S. pyogenes strain in mice and which could be used to identify key aspects of this bacteria's pathogenesis.
- Monocyte-derived dendritic cells early exposed to Mycobacterium tuberculosis induce an enhanced T helper 17 response and transfer mycobacterial antigens. [JOURNAL ARTICLE]
- Int J Med Microbiol 2016 Jun 23.
Tuberculosis (TB) is a complex disease, and the success of the bacterium depends on its ability to evade the immune response. Previously, we determined that Mycobacterium tuberculosis (Mtb) impairs the function of dendritic cells (DC), promoting the generation of cells that are poor stimulators of mycobacterial antigen-specific CD4T cells, which are required to control this persistent infection. In this study, we aimed to determine the mechanisms by which monocyte-derived DCs differentiated in the presence of Mtb (MtbDC) may impact on the proliferation of specific anti-mycobacterial T cells. We found that the presence of Mtb during monocyte-derived DC differentiation favours T helper (Th) 2 and Th17 polarization, in detriment of a Th1 response, compared to DC mature with Mtb. The bias on T cell polarization was associated to the profile of C-type lectin receptors expression found in MtbDC (DC-SIGN(low)/MR(low)/Dectin-1(high)). Alternatively, MtbDC release Mtb antigens (Ag) that can be taken up and presented by bystander DC, promoting the proliferation of CD4T cells, but to a lesser extent than direct presentation by Mtb-matured DC. In summary, we have further characterized the generation of MtbDC as an effective evasion strategy driven by the pathogen, leading to the inhibition of Ag-presentation and bias of T cell polarization towards Th2 and Th17 profiles, features which partially explain the persistence of Mtb in the host.