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Frontiers in Microbiology 2019is a commensal colonizer of both humans and animals, but also an opportunistic pathogen responsible for a multitude of diseases. In recent years, colonization of pigs...
is a commensal colonizer of both humans and animals, but also an opportunistic pathogen responsible for a multitude of diseases. In recent years, colonization of pigs by methicillin resistant has become a problem with increasing numbers of humans being infected by livestock strains. In colonization and virulence factor expression is controlled by the quorum sensing system, which responds to and is activated by self-generated, autoinducing peptides (AIPs). AIPs are also produced by coagulase negative staphylococci (CoNS) commonly found as commensals in both humans and animals, and interestingly, some of these inhibit activity. Here, we have addressed if cross-communication occurs between and CoNS strains isolated from pig nares, and if so, how properties such as host factor binding and biofilm formation are affected. From 25 pig nasal swabs we obtained 54 staphylococcal CoNS isolates belonging to 8 different species. Of these, none were able to induce as monitored by reporter gene fusions to regulated genes but a number of -inhibiting species were identified including , , , , and . After establishing that the inhibitory activity was mediated via AgrC, the receptor of AIPs, we synthesized selective AIPs to explore their effect on adhesion of to fibronectin, a host factor involved in colonization. Here, we found that the CoNS AIPs did not affect adhesion of except for strain 8325-4. When individual CoNS strains were co-cultured together with we observed variable degrees of biofilm formation which did not correlate with interactions. Our results show that multiple CoNS species can be isolated from pig nares and that the majority of these produce AIPs that inhibit . Further they show that the consequences of the interactions between CoNS and are complex and highly strain dependent.
PubMed: 31611856
DOI: 10.3389/fmicb.2019.02212 -
Journal of Dairy Science Dec 2019Coagulase-negative staphylococci (CNS) are one of the most common bovine mastitis pathogens found worldwide. In this study, we investigated the prevalence and...
Coagulase-negative staphylococci (CNS) are one of the most common bovine mastitis pathogens found worldwide. In this study, we investigated the prevalence and distribution of CNS species in mastitis milk samples and further characterized the methicillin-resistant (MR) CNS. A total of 311 CNS were isolated from 3,692 quarter milk samples from 1,373 dairy cattle at 81 farms between 2013 and 2017. Further evaluation of the CNS isolates revealed 14 CNS species among the samples and 3 predominant species-namely, Staphylococcus chromogenes, Staphylococcus simulans, and Staphylococcus epidermidis. Resistance was higher in S. epidermidis than in other CNS species except for resistance against oxacillin in Staphylococcus sciuri. Resistance to β-lactams was the most common in all CNS species (8.4% in ampicillin, 21.2% in oxacillin, and 13.5% in penicillin). Conversely, only minimal resistance to cephalothin, ceftiofur, and pirlimycin/novobiocin was found. Twenty-one isolates from 4 species were mecA-carrying MRCNS strains, including 18 S. epidermidis and 1 each of S. sciuri, Staphylococcus equorum, and Staphylococcus hominis. The majority of the mecA-carrying MRCNS isolates were produced in the biofilm. Furthermore, multidrug-resistant sequence type 179 isolate produced the strongest biofilm. Seven genotypes were detected in the 18 MR S. epidermidis strains, the most predominant of which persisted on a farm for 2 yr. Our findings for the antimicrobial susceptibility profiles and genotypic characterization of the MRCNS isolates could provide valuable information for controlling the spread of resistance and the selection of appropriate antimicrobial therapies for mastitis in the future. Further, strategic antibiotic use for mastitis treatment and hygienic management practices aimed at the prevention of the growth of resistant bacteria are urgently needed on dairy farms.
Topics: Animals; Anti-Bacterial Agents; Cattle; Coagulase; Drug Resistance, Bacterial; Farms; Female; Genotype; Mastitis, Bovine; Milk; Oxacillin; Prevalence; Republic of Korea; Staphylococcal Infections; Staphylococcus; Staphylococcus epidermidis
PubMed: 31548061
DOI: 10.3168/jds.2019-17028 -
ACS Applied Bio Materials Aug 2019This study presents a scalable method for designing magnetic Janus nanoparticles, which are capable of performing bacterial capture, while preventing agglomeration...
This study presents a scalable method for designing magnetic Janus nanoparticles, which are capable of performing bacterial capture, while preventing agglomeration between bacterial cells. To this end, we prepared silica-coated magnetite Janus nanoparticles functionalized with a bacteria-specific antibody on one side and polyethylene glycol chains on the other, using the established wax-in-water emulsion strategy. These magnetic Janus nanoparticles specifically interact with one type of bacteria from a mixture of bacteria via specific antigen-antibody interactions. Contrarily to bacterial capture with isotropically functionalized particles, the bacterial suspensions remain free from cell-nanoparticle-cell agglomerates, owing to the passivation coating with polyethylene glycol chains attached to the half of the magnetic nanoparticles pointing away from the bacterial surface after capture. The selective magnetic capture of cells was achieved from a mixture with without compromising bacterial viability and with an efficiency over 80%. This approach is a promising method for rapid and agglomeration-free separation of live bacteria for identification, enrichment, and cell counting of bacteria from biological samples.
PubMed: 35030739
DOI: 10.1021/acsabm.9b00415 -
AIMS Microbiology 2019The increasing spread of antibiotic-resistant microorganisms has led to the necessity of developing alternative antimicrobial treatments. The use of peptidoglycan...
The increasing spread of antibiotic-resistant microorganisms has led to the necessity of developing alternative antimicrobial treatments. The use of peptidoglycan hydrolases is a promising approach to combat bacterial infections. In our study, we constructed a 2 kb-triple-acting fusion gene () encoding the N-terminal amidase-5 domain of streptococcal LambdaSA2 prophage endolysin (D-glutamine-L-lysin endopeptidase), a mid-protein amidase-2 domain derived from the staphylococcal phage 2638A endolysin (N-acetylmuramoyl-L-alanine amidase) and the mature version (246 residues) of the Lysostaphin bacteriocin (glycyl-glycine endopeptidase) at the C-terminus. The gene was expressed in plants using the non-replicating (CPMV)-based vector pEAQ-HT and the replicating AltMV)-based pGD5TGB123-MCS-CP3 vector, and in using pET expression vectors pET26b+ and pET28a+. The resulting poor expression of this fusion protein in plants prompted the construction of a gene codon-optimized for expression in tobacco plants, resulting in an improved codon adaptation index (CAI) from 0.79 ( gene) to 0.93 ( gene). Incorporation of the nt gene into the pEAQ-HT vector, followed by transient expression in , led to accumulation of TFnt to an approximate level of 0.12 mg/g of fresh leaf weight. Antimicrobial activity of purified plant- and bacterial-produced TFnt proteins was assessed against two strains of Gram-positive 305 and Newman. The results showed that plant-produced TFnt protein was preferentially active against 305, showing 14% of growth inhibition, while the bacterial-produced TFnt revealed significant antimicrobial activity against both strains, showing 68 (IC 25 µg/ml) and 60% (IC 71 µg/ml) growth inhibition against 305 and Newman, respectively. Although the combination of codon optimization and transient expression using the non-replicating pEAQ-HT expression vector facilitated production of the TFnt protein in plants, the most functionally active antimicrobial protein was obtained using the prokaryotic expression system.
PubMed: 31384710
DOI: 10.3934/microbiol.2019.2.158 -
Wound Repair and Regeneration :... Nov 2019Diabetic foot ulcers (DFUs) are a major clinical problem exacerbated by prolonged bacterial infection. Macrophages, the primary innate immune cells, are multifunctional...
Diabetic foot ulcers (DFUs) are a major clinical problem exacerbated by prolonged bacterial infection. Macrophages, the primary innate immune cells, are multifunctional cells that regulate diverse processes throughout multiple phases of wound healing. To better understand the influence of microbial species on macrophage behavior, we cultured primary human monocyte-derived macrophages from four donors for 24 hours in media conditioned by bacteria and fungi (Pseudomonas aeruginosa, Corynebacterium amycolatum, Corynebacterium striatum, Staphylococcus aureus, Staphylococcus simulans, and Candida albicans) isolated from the DFUs of six patients. The effects of these microbe-derived signals on macrophage behavior were assessed by measuring the gene expression of a panel of 25 genes related to macrophage phenotype, angiogenesis, bacterial recognition, and cell survival, as well as secretion of two inflammatory cytokines using NanoString multiplex analysis. Principal component analysis showed that macrophage gene expression and protein secretion were affected by both microbial species as well as human donor. S. simulans and C. albicans caused up-regulation of genes associated with a proinflammatory (M1) phenotype, and P. aeruginosa caused an increase in the secretion of the proinflammatory cytokine and M1 marker tumor necrosis factor-alpha (TNFα). Together, these results suggest that macrophages respond to secreted factors from microbes by up-regulating inflammatory markers, and that the effects are strongly dependent on the monocyte donor. Ultimately, increased understanding of macrophage-microbe interactions will lead to the development of more targeted therapies for DFU healing.
Topics: Adult; Aged; Anti-Infective Agents; Bacterial Infections; Biopsy, Needle; Cells, Cultured; Cohort Studies; Culture Media; Diabetic Foot; Enzyme-Linked Immunosorbent Assay; Female; Humans; Macrophages; Male; Microbiota; Middle Aged; Sensitivity and Specificity; Wound Healing
PubMed: 31343792
DOI: 10.1111/wrr.12752 -
Foodborne Pathogens and Disease Oct 2019Bulk tank milk from 58 dairy goat and sheep flocks located in southern Spain was examined to determine the prevalence and distribution of . A total of 45 isolates were...
Bulk tank milk from 58 dairy goat and sheep flocks located in southern Spain was examined to determine the prevalence and distribution of . A total of 45 isolates were obtained and characterized to determine the species, antimicrobial resistance profile, and genetic similitude by pulse-field gel electrophoresis (PFGE) using I. isolates were confirmed by polymerase chain reaction (PCR) analysis of , and resistance to methicillin was determined by PCR analysis of A. A total of 10 different staphylococcal species were identified, 22.2% and 77.8% of which were coagulase positive and negative, respectively. Twenty-two (48.89%) isolates were resistant to at least one antimicrobial agent. Higher antimicrobial resistance values were obtained against tetracycline (28.9%) and penicillin (22.2%). Two isolates ( and ) were resistant to cefoxitin; however, none of the 45 isolates harbored A. Thirty pulsotypes were detected by PFGE. Interestingly, some isolates of , , , and showed high genetic similarity (>80%). These data suggest that genetically similar staphylococcal isolates circulate among goat and sheep dairy herds, and their different resistance patterns could be influenced by the management systems used.
Topics: Animals; Anti-Bacterial Agents; Drug Resistance, Bacterial; Female; Goats; Microbial Sensitivity Tests; Milk; Polymerase Chain Reaction; Prevalence; Sheep; Spain; Staphylococcal Infections; Staphylococcus; Staphylococcus aureus
PubMed: 31157980
DOI: 10.1089/fpd.2018.2593 -
The Journal of Dairy Research May 2019The objectives of the research described here were to describe the persistence of intramammary infections (IMI) caused by coagulase negative staphylococci (CNS) in goats...
The objectives of the research described here were to describe the persistence of intramammary infections (IMI) caused by coagulase negative staphylococci (CNS) in goats using strain-typing, and to evaluate the relationship between species-specific CNS IMI and somatic cell score (SCS) at the udder-half level. Udder-half milk samples were collected from all 909 lactating goats (1817 halves; 1 blind half) in a single herd. Milk samples were cultured on Columbia blood agar, and 220 goats with at least one half yielding a single colony type CNS were enrolled for two additional half-level samplings at approximately 1-month intervals. Isolates were identified to the species level by matrix-assisted laser desorption-ionisation time-of-flight mass spectrometry or PCR amplification and partial sequencing of tuf or rpoB. An IMI was defined as persistent when ≥1 follow-up sample yielded the same species and strain as on Day 0 based on pulsed-field gel electrophoresis. A generalised mixed linear model was used to evaluate the odds of persistence as a function of CNS species. A mixed linear model was used to evaluate the relationship between IMI status on a given day and SCS. Among 192 IMI, 69.8% were persistent based on species and strain-type. Staphylococcus simulans IMI had higher odds of persistence than Staphylococcus arlettae IMI. In primiparous goats, Staphylococcus epidermidis IMI was associated with higher SCS than S. arlettae, Staphylococcus xylosus and 'other CNS' IMI. The differences detected in the present study between CNS species, with regard to persistence of IMI and association with SCS, highlight the need to study CNS at the species and strain level to understand the pathogenicity and epidemiology of CNS in goats.
Topics: Animals; Cross-Sectional Studies; Female; Goat Diseases; Goats; Mastitis; Milk; Staphylococcal Infections; Staphylococcus
PubMed: 31138341
DOI: 10.1017/S0022029919000311 -
International Journal of Systematic and... Aug 2019A novel type strain, designated SDB 2975 (=CECT 9737=DSM 105892), of the novel species Staphylococcus debuckii sp. nov. isolated from bovine milk is described. The novel...
A novel type strain, designated SDB 2975 (=CECT 9737=DSM 105892), of the novel species Staphylococcus debuckii sp. nov. isolated from bovine milk is described. The novel species belongs to the genus Staphylococcus and showed resistance to tetracycline and was oxidase- and coagulase-negative, catalase-positive, and Gram-stain-positive. Phylogenetic relationships of Staphylococcus debuckii SDB 2975 to other staphylococcal species were inferred from 16S rRNA gene and whole-genome-based phylogenetic reconstruction. The 16S rRNA gene comparisons showed that the strain is closely related to Staphylococcus condimenti (99.73 %), Staphylococcus piscifermentans (99.66 %), Staphylococcus carnosus (99.59 %) and Staphylococcus simulans (98.03 %). Average nucleotide identity (ANI) values between S.taphylococcus debuckii SDB 2975 and its closely related Staphylococcus species were 83.96, 94.5, 84.03 and 78.09 %, respectively, and digital DNA-DNA hybridization (dDDH) values were 27.70, 58.02, 27.70 and 22.00 %, respectively. The genome of Staphylococcus debuckii SDB 2975 was sequenced with PacBio and Illumina technologies and is 2 691 850 bp long, has a G+C content of 36.6 mol% and contains 2678 genes and 80 RNAs, including six copies of each5S rRNA, 16S rRNA and 23S rRNA genes. Biochemical profiling and a newly developed PCR assay enabled differentiation of Staphylococcus debuckii SDB 2975 and three other SDB strains from its closest staphylococcal species. Differentiation was also achieved by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF). Genes unique to Staphylococcus debuckii were identified and a PCR-based assay was developed to differentiate Staphylococcus debuckii from other staphylococcal species. In conclusion, the results of phylogenetic analysis along with the ANI values <95 %, and dDDH values <70 % from closely related species along with the phenotypic and biochemical characteristics and specific MALDI-TOF profiles demonstrated that Staphylococcus debuckii SDB 2975 represents a novel species within the genus Staphylococcus, named Staphylococcus debuckii sp. nov. (SDB 2975=CECT 9737=DSM 105892).
Topics: Animals; Bacterial Typing Techniques; Base Composition; Cattle; DNA, Bacterial; Fatty Acids; Milk; Nucleic Acid Hybridization; Phylogeny; Quebec; RNA, Ribosomal, 16S; Sequence Analysis, DNA; Staphylococcus
PubMed: 31135334
DOI: 10.1099/ijsem.0.003457 -
Frontiers in Microbiology 2019Biogenic amines (BAs) in sausages represent a health risk for consumers, and thus investigating the BAs accumulation mechanism is important to control the BAs. In this...
Biogenic amines (BAs) in sausages represent a health risk for consumers, and thus investigating the BAs accumulation mechanism is important to control the BAs. In this study, the BAs profiles of 16 typical Chinese sausage samples were evaluated, and 8 kinds of common BAs were detected from different samples. As a whole, the BAs contents of the majority of Chinese sausage samples were within the safe dosage range, except that the total BAs and histamine concentrations of sample HBBD were above the toxic dosage levels. Furthermore, the bacterial and fungal communities of the Chinese sausage samples were investigated by high-throughput sequencing analysis, and , , , , , and were identified as the predominant genera. Accordingly, 13 representative strains were selected from the dominant genera, and their BAs formation and degradation properties were evaluated. Finally, the results of fermented meats model experiment indicated that the isolates including Sp, Se, Sc1, Sc2, and Ss could significantly reduce BAs, possessing the potential as the starter cultures to control the BAs in fermented meat products. The present study not only helped to explain the BAs accumulation mechanism in Chinese sausage, but also developed the candidates for potential BAs control in fermented meat products.
PubMed: 31130922
DOI: 10.3389/fmicb.2019.00872 -
Foodborne Pathogens and Disease Aug 2019This study was undertaken to investigate the antimicrobial resistance patterns of major causative agents to clinical mastitis in Danish dairy cows collected in 2016 to...
This study was undertaken to investigate the antimicrobial resistance patterns of major causative agents to clinical mastitis in Danish dairy cows collected in 2016 to provide data on the current resistance patterns. Such data may subsequently serve as basis for a guideline for prudent use of antimicrobial agents in mastitis treatment. In addition, this study serves as a baseline for future comparison. The minimum inhibitory concentrations in ( = 62), ( = 18), ( = 63), coagulase-negative Staphylococci (CNS) ( = 49), ( = 61), ( = 33), and ( = 13) were determined to antimicrobial agents representing most classes relevant for treatment. The occurrence of resistance in the 299 bacterial isolates in total was evaluated using Clinical and Laboratory Standards Institute clinical breakpoints or in-house breakpoint values. For , low resistance levels were detected, 11.3% being resistant to ampicillin while resistance to other compounds was lower or zero. In contrast, revealed frequent ampicillin resistance (83.3%), but was susceptible to most other antimicrobial agents tested. Staphylococci were susceptible to the majority of antimicrobial agents tested, only 17.7% of the isolates and 22.4% of the CNS being resistant to penicillin. Species distribution of the CNS isolates revealed that , , and were the most prevalent species. One and one isolate was found to be cefoxitin resistant and confirmed as methicillin resistant by polymerase chain reaction detection of the gene, showing that methicillin resistance in staphylococci is present. All species of streptococci were susceptible to penicillin. No other critical resistance was found in any species, and resistance was in general low to all clinically relevant compounds. We emphasize the need for continuous surveillance of antibiotic resistance in major mastitis pathogens and the need for harmonization of methods and interpretations.
Topics: Animals; Anti-Bacterial Agents; Cattle; Dairying; Denmark; Drug Resistance, Bacterial; Escherichia coli; Female; Food Microbiology; Mastitis, Bovine; Microbial Sensitivity Tests; Staphylococcus; Streptococcus
PubMed: 31059284
DOI: 10.1089/fpd.2018.2560