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1.
Klin Lab Diagn ; 66(1): 42-44, 2021 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-33567172

RESUMO

1235 strains of Staphylococci isolated in a multidisciplinary children's clinic were analyzed. The species and antibiotic resistance of Golden and coagulase-negative Staphylococci were studied. The most frequently identified species were: S. aureus-36.06%, S. epidermidis-23.05%, S. haemolyticus-19.7%, S. hominis-14.03%. Phenotype methicillinsensitive strains had 48.9% of the allocated staphylococci, while metitillinrezistentnykh S. aureus was identified in 25.6%, and coagulase-negative staphylococci methicillinresistant- 63.2 per cent. The frequency of associated resistance to aminoglycosides, fluoroquinolones, macrolides and tetracyclines have metitillinresictant strains 92,7%, 78,3%, 83,4% and 52,05% respectively, resistant Staphylococcus and coagulase-negative staphylococci were similar. The minimum number of resistant strains was found in relation to daptomycin, no strains resistant to vancomycin and linezolid were found. Antibiotic resistance of staphylococci in children's hospitals is determined by the presence of the mecA gene or sensitivity to cefoxitin and does not depend on the type of strain.


Assuntos
Staphylococcus aureus Resistente à Meticilina , Infecções Estafilocócicas , Antibacterianos/farmacologia , Criança , Coagulase/genética , Resistência Microbiana a Medicamentos , Hospitais , Humanos , Testes de Sensibilidade Microbiana , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/epidemiologia , Staphylococcus/genética , Staphylococcus aureus
2.
Int J Food Microbiol ; 337: 108932, 2021 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-33152570

RESUMO

Culturing methods are conventionally applied to investigate the contamination of food with several microorganisms after heat processing. However, with these methods, it is not possible to evaluate whether heat-treated meat products, such as cooked sausages, contained parts of spoiled meat. Therefore, two specific multiplex qPCRs were developed in this study in order to determine the microbiological quality of the raw materials used for these products. The PCR targets focused on four bacterial groups often found on meat (family Enterobacteriaceae, genus Pseudomonas, genus Staphylococcus and species Brochothrix thermosphacta). Specificity as well as sensitivity of the developed multiplex qPCRs, validated by using 68 microbial species, were 100%. The applicability of both multiplex qPCRs compared to culturing methods was performed using 96 meat samples (fresh and naturally spoiled) and 12 inhouse-made "Lyoner" sausages containing variable ratios of spoiled meat (0%, 5%, 12% and 25%; n = 3 for each group). Both methods showed similar results by evaluating the ∆log10 cfu/g, the relative accuracy and the t-test analysis (p > 0.05). Comparing qPCR results of the different sausage groups, a significant difference between sausages containing fresh meat and sausages containing spoiled meat (12% and 25%) was found only for Pseudomonas and B. thermosphacta in both raw and cooked sausages. The statistical difference between 5% vs. 12% and 25% spoiled meat in cooked sausages, was also found only for these two bacterial groups. The developed multiplex qPCRs were further applied to 30 commercially available "Bologna-type" sausages. The results showed a total of 14 sausages considered to be suspicious for Food Fraud. While the role of Staphylococcus spp. in meat spoilage remains unclear, Pseudomonas, Enterobacteriaceae and B. thermosphacta could together be used as an indicator for "spoiled meat" used in sausages. The developed qPCR systems in this study allow the detection of four relevant bacterial groups in the heated Bologna-type sausages and provide information about the hygienic quality of raw materials used. This method could thus be helpful for screening food suspected of Food Fraud.


Assuntos
Bactérias/genética , Microbiologia de Alimentos/métodos , Produtos da Carne/microbiologia , Carne/microbiologia , Reação em Cadeia da Polimerase , Animais , Brochothrix/genética , Enterobacteriaceae/genética , Temperatura Alta , Pseudomonas/genética , Staphylococcus/genética
3.
Int J Food Microbiol ; 336: 108902, 2021 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-33091757

RESUMO

Routine evaluation of the slaughter process is performed by the enumeration of the aerobic colony count, Enterobacteriaceae and Salmonella spp. on the carcass through destructive or non-destructive methods. With non-destructive methods, bacteria are counted from a minimum area of 100 cm2 in different sampling sites on the pork carcasses, and the results of these investigated areas are pooled to one value for the complete carcass evaluation (a total of 400 cm2). However, the composition of the bacterial community present on the different sampling areas remains unknown. The aim of the study was to characterize the microbial population present on four areas (ham, back, jowl and belly) of eight pork carcasses belonging to two different slaughterhouses through culture-dependent (Matrix-assisted laser desorption/ionization time-of-flight Mass Spectrometry MALDI-TOF MS, combined with 16S rRNA gene sequencing) and complementary culture-independent (16S rRNA amplicon sequencing) methods. The presence of Salmonella spp. and Y. enterocolitica was additionally assessed. Using MALDI-TOF MS, Staphylococcus, Pseudomonas, and Escherichia coli were found to dominate the bacterial cultures isolated from the 8 carcasses. Based on the 16S rRNA amplicon sequencing analyses however, no specific genus clearly dominated the bacterial community composition. By using this culture-independent method, the most abundant genera in microbial populations of the ham, back, jowl and belly were found to be similar, but important differences between the two slaughterhouses were observed. Thus, present data suggests that the indigenous bacterial population of individual animals is overruled by the microbial population of the slaughterhouse in which the carcass is handled. Also, our data suggests that sampling of only one carcass area by official authorities may be appropriate for the evaluation of the hygienic status of the carcasses and therefore of the slaughter process.


Assuntos
Matadouros , Bactérias/genética , Microbiologia de Alimentos , Carne/microbiologia , Animais , Bactérias/classificação , Bactérias/isolamento & purificação , Enterobacteriaceae/isolamento & purificação , Escherichia coli/genética , Pseudomonas/genética , RNA Ribossômico 16S/genética , Salmonella/genética , Staphylococcus/genética , Suínos
4.
J Water Health ; 18(6): 890-898, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33328361

RESUMO

This study examined the occurrence and molecular basis for antibiotic-resistant staphylococci from the wastewater treatment plant and grey-water samples in Obafemi Awolowo University, Nigeria. Standard microbiological techniques and molecular methods were utilized. The species identified (MALDI score >1.7) comprised S. saprophyticus (19), S. cohnii (8), S. sciuri (7), S. aureus (4), S. epidermidis (3), S. warneri (2), S. equorum (1), S. haemolyticus (1), S. nepalensis (1), S. condimenti (1), and S. pasteuri (1). Resistance to trimethoprim, tetracycline and cefoxitin were observed in 78.3% (47/60), 36.7% (22/60) and 25% (15/60) of the isolates, respectively. The rate of multidrug resistance was 53.3% (32/60) and observed in eight species from different sampling sites. Seven (S. sciuri; n = 5; S. aureus; n = 1; S. warneri; n = 1) of the 20 selected (representing the various staphylococcal species and antibiotypes) isolates were mecA-positive. Furthermore, the tetK gene was detected in nine isolates, six with dfrA, and four were positive for the dfrG gene. One S. aureus was mecA, tetK and dfrG gene positive. The study provides insights on antibiotic-resistant staphylococci from a non-clinical setting and highlights the need for active surveillance to understand the burden of antimicrobial resistance in Nigeria. This is key to improve synergy across the human, animal and environmental health sectors in Nigeria.


Assuntos
Staphylococcus , Purificação da Água , Animais , Antibacterianos/farmacologia , Humanos , Testes de Sensibilidade Microbiana , Nigéria , Staphylococcus/genética , Staphylococcus aureus/genética , Universidades , Água
5.
An Acad Bras Cienc ; 92(3): e20180925, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33053101

RESUMO

In this work, we investigated the phenotypic profile of Staphylococcus spp. isolates recovered from raw milk and artisanal cheese, and their enterotoxigenic potential through the detection of classical enterotoxin genes (sea, seb, sec, sed and see). A total of 104 isolates (58 coagulase-positive Staphylococcus - CoPS; and 46 coagulase-negative Staphylococcus- CoNS) were used, of which 33 were retrieved from raw milk and 71 from artisanal cheese produced in the Serrana region of Santa Catarina. Identification of CoPS was conducted via biochemical tests. Detection of the genes sea, seb, sec, sed, and see was carried out by multiplex PCR technique. Among the 58 CoPS analyzed, 64% were identified as S. aureus, 22% as S. scheiferi coagulans, 12% as S. hyicus and as 2% S. intermedius. In the present study was noted that 40% of CoPS isolates retrieved from milk harbored seb gene, while only one from artisanal cheese was positive for gene sea. In this study all CoNS samples investigated were negative for enterotoxins genes. The enterotoxigenic potential of CoPS, is an issue of great importance for public health. For that reason, it is necessary that cheese factories strictly follow the safety processes involved in manufacturing.


Assuntos
Queijo , Staphylococcus , Animais , Microbiologia de Alimentos , Leite , Staphylococcus/genética , Staphylococcus/isolamento & purificação , Staphylococcus aureus
6.
Nat Commun ; 11(1): 5431, 2020 10 27.
Artigo em Inglês | MEDLINE | ID: mdl-33110079

RESUMO

Physical forces have profound effects on cellular behavior, physiology, and disease. Perhaps the most intruiguing and fascinating example is the formation of catch-bonds that strengthen cellular adhesion under shear stresses. Today mannose-binding by the Escherichia coli FimH adhesin remains one of the rare microbial catch-bond thoroughly characterized at the molecular level. Here we provide a quantitative demonstration of a catch-bond in living Gram-positive pathogens using force-clamp spectroscopy. We show that the dock, lock, and latch interaction between staphylococcal surface protein SpsD and fibrinogen is strong, and exhibits an unusual catch-slip transition. The bond lifetime first grows with force, but ultimately decreases to behave as a slip bond beyond a critical force (~1 nN) that is orders of magnitude higher than for previously investigated complexes. This catch-bond, never reported for a staphylococcal adhesin, provides the pathogen with a mechanism to tightly control its adhesive function during colonization and infection.


Assuntos
Adesinas Bacterianas/química , Infecções Estafilocócicas/microbiologia , Staphylococcus/metabolismo , Adesinas Bacterianas/genética , Adesinas Bacterianas/metabolismo , Aderência Bacteriana , Fibrinogênio/química , Fibrinogênio/metabolismo , Humanos , Ligação Proteica , Análise Espectral , Infecções Estafilocócicas/metabolismo , Staphylococcus/química , Staphylococcus/genética , Staphylococcus/crescimento & desenvolvimento
7.
Lett Appl Microbiol ; 71(4): 359-368, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32713031

RESUMO

Staphylococci from Sheedal of Northeast India was isolated, identified and characterized. All the isolated staphylococci were found to be coagulase negative. Based on the rpoB gene sequences followed by analysis using NCBI-BLAST software, seven species of Staphylococcus namely, S. piscifermentans, S. condimenti, S. arlettae, S. sciuri, S. warneri, S. nepalensis and S. hominis were recognized. Phylogenetic analyses revealed three major cluster groups. All the seven Staphylococcus showed their NaCl tolerance from 2 to 8%. No species was able to grow at 55°C. Except S. arlettae and S. sciuri, all the isolated staphylococcal species exhibited growth at pH 4-8. No isolated species was able to ferment mannitol, sucrose and arabinose. All the species exhibited moderate to maximum proteolytic and lipolytic activities. All the seven species were found to be sensitive to the antibiotics, namely, erythromycin, norfloxacin, ampicillin, streptomycin and vancomycin, whereas all were resistant to co-trimoxazole. Only S. piscifermentans was found antagonist to Salmonella enterica, Escherichia coli and Bacillus subtilis, although the clear zone was minimal. All the staphylococcal species except S. arlettae and S. sciuri exhibited hydrophobicity ranging from 25 to 66%. The observed characteristics of isolated Staphylococci from Sheedal revealed their role in fish fermentation.


Assuntos
Alimentos e Bebidas Fermentados/microbiologia , Produtos Pesqueiros/microbiologia , Staphylococcus/isolamento & purificação , Ampicilina/farmacologia , Animais , Antibacterianos/farmacologia , Eritromicina/farmacologia , Fermentação , Peixes/microbiologia , Contaminação de Alimentos/análise , Índia , Filogenia , Staphylococcus/classificação , Staphylococcus/efeitos dos fármacos , Staphylococcus/genética
8.
PLoS Pathog ; 16(6): e1007806, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32497104

RESUMO

Coagulase-positive staphylococci, which frequently colonize the mucosal surfaces of animals, also cause a spectrum of opportunistic infections including skin and soft tissue infections, urinary tract infections, pneumonia, and bacteremia. However, recent advances in bacterial identification have revealed that these common veterinary pathogens are in fact zoonoses that cause serious infections in human patients. The global spread of multidrug-resistant zoonotic staphylococci, in particular the emergence of methicillin-resistant organisms, is now a serious threat to both animal and human welfare. Accordingly, new therapeutic targets that can be exploited to combat staphylococcal infections are urgently needed. Enzymes of the methylerythritol phosphate pathway (MEP) of isoprenoid biosynthesis represent potential targets for treating zoonotic staphylococci. Here we demonstrate that fosmidomycin (FSM) inhibits the first step of the isoprenoid biosynthetic pathway catalyzed by deoxyxylulose phosphate reductoisomerase (DXR) in staphylococci. In addition, we have both enzymatically and structurally determined the mechanism by which FSM elicits its effect. Using a forward genetic screen, the glycerol-3-phosphate transporter GlpT that facilitates FSM uptake was identified in two zoonotic staphylococci, Staphylococcus schleiferi and Staphylococcus pseudintermedius. A series of lipophilic ester prodrugs (termed MEPicides) structurally related to FSM were synthesized, and data indicate that the presence of the prodrug moiety not only substantially increased potency of the inhibitors against staphylococci but also bypassed the need for GlpT-mediated cellular transport. Collectively, our data indicate that the prodrug MEPicides selectively and robustly inhibit DXR in zoonotic staphylococci, and further, that DXR represents a promising, druggable target for future development.


Assuntos
Antibacterianos , Farmacorresistência Bacteriana Múltipla , Pró-Fármacos , Infecções Estafilocócicas , Staphylococcus , Zoonoses , Animais , Antibacterianos/química , Antibacterianos/farmacologia , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Farmacorresistência Bacteriana Múltipla/genética , Humanos , Pró-Fármacos/química , Pró-Fármacos/farmacologia , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/genética , Infecções Estafilocócicas/metabolismo , Staphylococcus/genética , Staphylococcus/crescimento & desenvolvimento , Zoonoses/tratamento farmacológico , Zoonoses/genética , Zoonoses/metabolismo , Zoonoses/microbiologia
9.
Lett Appl Microbiol ; 71(4): 394-399, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32557652

RESUMO

The aims of the present study were to examine the occurrence of Staphylococcus spp. in the tonsils of slaughtered pigs in a regional slaughterhouse in Greece, the antibiotic resistance of the Staphylococcus spp. isolates, and the enteroxigenicity of the S. aureus isolates. Staphylococcus spp. were isolated in 70 (48·61%) out of the total 144 tonsil samples. The predominant species was S. aureus in coagulase-positive staphylococci (CoPS), while the predominant species were Staphylococcus epidermidis and Staphylococcus saprophyticus in the coagulase-negative staphylococci (CoNS). Staphylococcus spp. isolates presented high antibiotic resistance frequencies to tetracycline (97·1%) or clindamycin (80·0%) and low antibiotic resistance frequencies to fusidic acid (14·3%). No methicillin-resistant S. aureus (MRSA) strains were identified, and all Staphylococcus spp. isolates were susceptible to vancomycin. Among the 26 S. aureus isolates, 21 (80·76%) possessed staphylococcal enterotoxin genes with seven different enterotoxin gene profiles. The predominant enterotoxin profile was seg, sei and sej with seven S. aureus isolates. The occurrence of multidrug resistant Staphylococcus spp. in pig tonsils indicate public health risk to pork consumers and handlers in developing antimicrobial resistance.


Assuntos
Farmacorresistência Bacteriana , Enterotoxinas/metabolismo , Tonsila Palatina/microbiologia , Infecções Estafilocócicas/veterinária , Staphylococcus/isolamento & purificação , Doenças dos Suínos/microbiologia , Matadouros/estatística & dados numéricos , Animais , Antibacterianos/farmacologia , Clindamicina/farmacologia , Grécia/epidemiologia , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Infecções Estafilocócicas/epidemiologia , Infecções Estafilocócicas/microbiologia , Staphylococcus/classificação , Staphylococcus/efeitos dos fármacos , Staphylococcus/genética , Suínos , Doenças dos Suínos/epidemiologia
10.
Artigo em Inglês | MEDLINE | ID: mdl-32466464

RESUMO

Cell phones, smartphones, and tablets are extensively used in social and professional life, so they are frequently exposed to bacteria. The main goal of the present work was to isolate and characterize Staphylococci strains from students' cell phone mobiles. Subsequently, 24 Staphylococci strains were tested against a wide range of antibiotics, for the distribution of some virulence-related genes and their ability to form biofilm. Staphylococcus spp. were cultured from all studied devices on chromogenic medium and identified using the matrix-assisted laser desorption/ionization (MALDI), time-of-flight (TOF) mass spectrometry (MS) technique (MALDI-TOF-MS). The results obtained showed that S. aureus was the dominant species (19 strains, 79.1%), followed by S. warneri (3 strains, 12.5%), and S. haemolyticus (2 strains, 8.3%). Isolated strains showed high percentages of hydrolytic enzymes production, resistance to many tested antibiotics, and 37.5% expressed the mecA gene. The tested strains were highly adhesive to polystyrene and glass and expressed implicated icaA (62.5%) and icaD (66.6%) genes. All Staphylococcus spp. strains tested were found to possess proteases and the α-hemolysin gene. Our results highlighted the importance of mobile phones as a great source of Staphylococcus spp., and these species were found to be resistant to many antibiotics with multiple antibiotic resistance (MAR) index ranging from (0.444) to (0.812). Most of the studied strains are able to form biofilm and expressed many virulence genes. Phylogenetic analysis based on the phenotypic and genetic characters highlighted the phenotypic and genetic heterogeneity of the S. aureus population studied. Further analyses are needed to elucidate the human health risks associated with the identified Staphylococci strains.


Assuntos
Telefone Celular , Staphylococcus , Antibacterianos , Biofilmes , Farmacorresistência Bacteriana , Genótipo , Humanos , Fenótipo , Filogenia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Staphylococcus/genética , Staphylococcus/isolamento & purificação
11.
PLoS One ; 15(5): e0233363, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32442199

RESUMO

Staphylococcus warneri is a coagulase-negative staphylococcus that is a normal inhabitant of the skin. It is also considered to be an opportunistic etiological agent causing significant infections in human and animals. Currently, relatively little attention has been paid to the genome biology of S. warneri pathogenicity and antibiotic resistance, which are emerging issues for this etiological agent with considerably clinical significance. In this study, we determined the complete genome sequence of S. warneri strain GD01 recovered from the sampled muscle abscess tissue of a diseased pig in South China. The genome of S. warneri is composed of a circular chromosome of 2,473,911 base pairs as well as eight plasmid sequences. Genome-wide metabolic reconstruction revealed 82 intact functional modules driving the catabolism of respiration and fermentation for energy production, uptake of distinct sugars as well as two-component regulatory systems. The evidence uncovered herein enables better understanding for metabolic potential and physiological traits of this etiological agent. The antibiotic susceptibility test demonstrated that S. warneri GD01 was resistant to penicillin, amoxicillin, ampicillin, cefalexin, vancomycin, and sulfisoxazole. The associations between antibiotic phenotypes and the related genotypes were identified to reveal the molecular basis conferring resistance to this pathogen. A number of genes coding for potential virulence factors were firstly depicted in the genome of S. warneri GD01, including adhesins, exoenzymes, capsule, and iron acquisition proteins. Our study provides a valuable genomic context of the genes/modules devoting to metabolism, antibiotic resistance, and virulence of S. warneri.


Assuntos
Antibacterianos/farmacologia , DNA Bacteriano , Staphylococcus/efeitos dos fármacos , Staphylococcus/genética , Animais , China , Testes de Sensibilidade Microbiana , Suínos , Sequenciamento Completo do Genoma
12.
Arch Microbiol ; 202(7): 1899-1906, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32448960

RESUMO

Endophytic bacteria have been reported to have symbiotic, mutualistic, commensalistic or trophobiotic relationships with various plant parts. As part of its adaptation, many endophytic organisms are known to exhibit properties with multiple beneficial effects to the plant system. Even though many bacterial genera have been identified to have endophytic association, isolation of those which were previously demonstrated well for human association is quite interesting. In the study, endophytic bacteria Ceb1 isolated from the rhizome of Curcuma longa was identified by 16S rDNA sequencing as Staphylococcus sp. Further, Ceb1 was observed to have the ability to tolerate drought stress. While screening for the plant growth-promoting traits, Ceb1 was found to be positive for IAA production both under drought-stressed and normal conditions as confirmed by HPLC. The Ceb1 priming with Vigna unguiculata was observed to enhance the growth parameters of the plant. Analysis of Ceb1-treated plants by ICP-MS further showed modulation of both macro- and micronutrients. Upon drought stress induction in Vigna unguiculata, Ceb1 was found to provide synergistic plant growth-promoting effect to the plant along with the supplemented silicate sources. Under the changing agroclimatic conditions, exploring the plant stress-alleviating effects of endophytes is highly significant.


Assuntos
Secas , Desenvolvimento Vegetal , Plantas/microbiologia , Silicatos/farmacologia , Staphylococcus/fisiologia , Curcuma/microbiologia , Endófitos/classificação , Endófitos/genética , Endófitos/isolamento & purificação , Endófitos/metabolismo , Desenvolvimento Vegetal/fisiologia , RNA Ribossômico 16S/genética , Rizoma/microbiologia , Staphylococcus/genética , Staphylococcus/isolamento & purificação , Simbiose , Vigna/fisiologia
13.
PLoS Biol ; 18(3): e3000686, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32226015

RESUMO

Compact CRISPR/Cas9 systems that can be packaged into an adeno-associated virus (AAV) hold great promise for gene therapy. Unfortunately, currently available small Cas9 nucleases either display low activity or require a long protospacer adjacent motif (PAM) sequence, limiting their extensive applications. Here, we screened a panel of Cas9 nucleases and identified a small Cas9 ortholog from Staphylococcus auricularis (SauriCas9), which recognizes a simple NNGG PAM, displays high activity for genome editing, and is compact enough to be packaged into an AAV for genome editing. Moreover, the conversion of adenine and cytosine bases can be achieved by fusing SauriCas9 to the cytidine and adenine deaminase. Therefore, SauriCas9 holds great potential for both basic research and clinical applications.


Assuntos
Proteína 9 Associada à CRISPR/metabolismo , DNA/metabolismo , Edição de Genes/métodos , Staphylococcus/enzimologia , Sequência de Aminoácidos , Proteína 9 Associada à CRISPR/química , Proteína 9 Associada à CRISPR/genética , Sistemas CRISPR-Cas , DNA/química , DNA/genética , Dependovirus/genética , Células HEK293 , Humanos , Motivos de Nucleotídeos , Engenharia de Proteínas , Staphylococcus/genética , Especificidade por Substrato
14.
Appl Environ Microbiol ; 86(12)2020 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-32245765

RESUMO

Staphylococcus agnetis has been previously associated with subclinical or clinically mild cases of mastitis in dairy cattle and is one of several staphylococcal species that have been isolated from the bones and blood of lame broilers. We reported that S. agnetis could be obtained frequently from bacterial chondronecrosis with osteomyelitis (BCO) lesions of lame broilers (A. Al-Rubaye et al., PLoS One 10:e0143336, 2015 [https://doi.org/10.1371/journal.pone.0143336]). A particular isolate, S. agnetis 908, can induce lameness in over 50% of exposed chickens, exceeding normal BCO incidences in broiler operations. We reported the assembly and annotation of the genome of isolate 908. To better understand the relationship between dairy cattle and broiler isolates, we assembled 11 additional genomes for S. agnetis isolates, an additional chicken BCO strain, and ten isolates from cattle milk, mammary gland secretions, or udder skin from the collection at the University of Missouri. To trace phylogenetic relationships, we constructed phylogenetic trees based on multilocus sequence typing and genome-to-genome distance comparisons. Chicken isolate 908 clustered with two of the cattle isolates, along with three isolates from chickens in Denmark and an isolate of S. agnetis we isolated from a BCO lesion on a commercial broiler farm in Arkansas. We used a number of BLAST tools to compare the chicken isolates to those from cattle and identified 98 coding sequences distinguishing isolate 908 from the cattle isolates. None of the identified genes explain the differences in host or tissue tropism. These analyses are critical to understanding how staphylococci colonize and infect different hosts and potentially how they can transition to alternative niches (bone versus dermis).IMPORTANCE Staphylococcus agnetis has been recently recognized as associated with disease in dairy cattle and meat-type chickens. The infections appear to be limited in cattle and systemic in broilers. This report details the molecular relationships between cattle and chicken isolates in order to understand how this recently recognized species infects different hosts with different disease manifestations. The data show that the chicken and cattle isolates are very closely related, but the chicken isolates all cluster together, suggesting a single jump from cattle to chickens.


Assuntos
Bovinos/microbiologia , Galinhas/microbiologia , Genoma Bacteriano , Staphylococcus/classificação , Staphylococcus/genética , Animais , Filogenia , Staphylococcus/patogenicidade , Virulência/genética
15.
Balkan Med J ; 37(4): 215-221, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32270947

RESUMO

Background: Coagulase-negative staphylococci, which belong to the normal microbiota of the skin and mucous membranes, are opportunistic pathogens. sasX, a newly described protein, is thought to play an important role in nasal colonization and methicillin-resistant Staphylococcus aureus virulence, and it may be acquired from coagulase-negative staphylococci by horizontal gene transfer. It has been considered that understanding the function of sasX gene may help clarify the relevance of the different adhesion mechanisms in the pathogenesis of infections associated with biofilm. Aims: To investigate the sasX gene presence, staphylococcal cassette chromosome mec types, and antimicrobial resistance patterns of invasive and noninvasive coagulase-negative staphylococci isolates. Study Design: Cross-sectional study. Methods: The study included a total of 180 coagulase-negative staphylococci strains. Non-invasive isolates (n=91) were obtained from the hands of healthy volunteers who do not work at the hospital (n=30), the nasal vestibule of healthy volunteer hospital workers (n=26), and central venous catheter (n=35). Invasive isolates (n=89) were isolated from peripheral blood cultures of inpatients who do not have catheters. All isolates were identified by conventional microbiological methods, automated systems, and, if needed, with matrix-assisted laser desorption/ionization-time of flight. Staphylococcal cassette chromosome mec typing, sasX and mec gene detection, antibiotic susceptibility, and sasX gene sequence analysis were performed. Results: Peripheral blood, central venous catheter colonization, and nasal vestibule isolates were positive for the sasX gene, whereas hand isolates were negative. sasX gene was present in 17 isolates, and no statistical significance was found between invasive and noninvasive isolates (p=0.173). Sequence analysis of the sasX genes showed high homology to related proteins of Staphylococcus phage SPbeta-like and Staphylococcus epidermidis RP62A. staphylococcal cassette chromosome mec type V was the most prevalent regardless of species. staphylococcal cassette chromosome mec type II was more frequent in invasive isolates and found to be statistically important for invasive and noninvasive S. epidermidis isolates (p=0.029). Staphylococcus haemolyticus isolates had the overall highest resistance rates. Resistance to ciprofloxacin, trimethoprim-sulfamethoxazole, and erythromycin was found to be higher in isolates from catheter and blood culture. Staphylococcus hominis isolates had the highest rate for inducible clindamycin resistance. None of the isolates were resistant to vancomycin, teicoplanin, and linezolid. Conclusion: The sasX gene is detected in 9.44% of the isolates. There is no statistical difference between the sasX-positive and -negative isolates in terms of antibacterial resistance and the presence of sasX and SCCmec types. Further studies about the role of sasX at virulence in coagulase-negative staphylococci, especially from clinical samples such as tracheal aspirate and abscess isolates, and distribution of staphylococcal cassette chromosome mec types are needed.


Assuntos
Coagulase/análise , Staphylococcus/genética , Staphylococcus/metabolismo , Coagulase/sangue , Coagulase/metabolismo , Estudos Transversais , Humanos , Testes de Sensibilidade Microbiana/métodos , Staphylococcus/isolamento & purificação , Staphylococcus capitis/genética , Staphylococcus capitis/isolamento & purificação , Staphylococcus epidermidis/genética , Staphylococcus epidermidis/isolamento & purificação , Staphylococcus haemolyticus/genética , Staphylococcus haemolyticus/isolamento & purificação , Staphylococcus hominis/genética , Staphylococcus hominis/isolamento & purificação , Staphylococcus lugdunensis/genética , Staphylococcus lugdunensis/isolamento & purificação , Staphylococcus saprophyticus/genética , Staphylococcus saprophyticus/isolamento & purificação
16.
Vet Microbiol ; 243: 108631, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32273010

RESUMO

This work aimed at characterizing four Staphylococcus aureus and 68 coagulase-negative staphylococci (CoNS), recovered from the air and liquid manure tank of two swine farms with intensive- and semi-extensive-production types, for their antimicrobial resistance pheno-/genotypes and their virulence gene content. Molecular typing was performed by spa typing, MLST, agr typing, and SCCmec typing, where applicable. Conjugation experiments were performed to assess the transferability of the linezolid resistance gene cfr, and its genetic environment was determined by Whole-Genome-Sequencing. The four S. aureus (intensive-production farm, IP-farm) were typed as t011-agrI-CC398-ST398, were scn-negative and two of them were methicillin-resistant (MRSA) with the mecA gene (SCCmec-V). Multidrug resistance was seen in 87 % of the CoNS. Statistically significant differences among the antimicrobial resistance rates of CoNS from the two farms were observed for cefoxitin, aminoglycosides, tetracycline, ciprofloxacin and trimethoprim-sulfamethoxazole. Eight methicillin-resistant CoNS, which were recovered from the IP-farm, carried the mecA gene. One S. simulans isolate was PVL-positive and three S. cohnii eta-positive. One S. equorum and one S. arlettae showed linezolid resistance and carried the cfr gene (IP-farm), which was non-transferable by conjugation into S. aureus. The cfr genetic context in both isolates was identical, with the lsa(B) gene located upstream of cfr. The environment of swine farms might contribute to the dissemination of CoNS that show multidrug resistance and harbor important virulence factors.


Assuntos
Proteínas de Bactérias/genética , Toxinas Bacterianas/genética , Exotoxinas/genética , Leucocidinas/genética , Infecções Estafilocócicas/veterinária , Staphylococcus/genética , Microbiologia do Ar , Animais , Antibacterianos/farmacologia , Técnicas de Tipagem Bacteriana , Coagulase , Farmacorresistência Bacteriana Múltipla/genética , Fazendas , Genes Bacterianos , Esterco/microbiologia , Meticilina/farmacologia , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/genética , Testes de Sensibilidade Microbiana , Tipagem de Sequências Multilocus , Staphylococcus/efeitos dos fármacos , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/genética , Suínos , Fatores de Virulência/genética
17.
Proc Natl Acad Sci U S A ; 117(12): 6531-6539, 2020 03 24.
Artigo em Inglês | MEDLINE | ID: mdl-32156733

RESUMO

Anti-CRISPRs (Acrs) are small proteins that inhibit the RNA-guided DNA targeting activity of CRISPR-Cas enzymes. Encoded by bacteriophage and phage-derived bacterial genes, Acrs prevent CRISPR-mediated inhibition of phage infection and can also block CRISPR-Cas-mediated genome editing in eukaryotic cells. To identify Acrs capable of inhibiting Staphylococcus aureus Cas9 (SauCas9), an alternative to the most commonly used genome editing protein Streptococcus pyogenes Cas9 (SpyCas9), we used both self-targeting CRISPR screening and guilt-by-association genomic search strategies. Here we describe three potent inhibitors of SauCas9 that we name AcrIIA13, AcrIIA14, and AcrIIA15. These inhibitors share a conserved N-terminal sequence that is dispensable for DNA cleavage inhibition and have divergent C termini that are required in each case for inhibition of SauCas9-catalyzed DNA cleavage. In human cells, we observe robust inhibition of SauCas9-induced genome editing by AcrIIA13 and moderate inhibition by AcrIIA14 and AcrIIA15. We also find that the conserved N-terminal domain of AcrIIA13-AcrIIA15 binds to an inverted repeat sequence in the promoter of these Acr genes, consistent with its predicted helix-turn-helix DNA binding structure. These data demonstrate an effective strategy for Acr discovery and establish AcrIIA13-AcrIIA15 as unique bifunctional inhibitors of SauCas9.


Assuntos
Proteínas de Bactérias/metabolismo , Proteína 9 Associada à CRISPR/antagonistas & inibidores , Sistemas CRISPR-Cas , Inibidores Enzimáticos/metabolismo , Staphylococcus/genética , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Proteína 9 Associada à CRISPR/genética , Proteína 9 Associada à CRISPR/metabolismo , Sistemas CRISPR-Cas/genética , Sequência Conservada , DNA/metabolismo , Edição de Genes , Genoma Bacteriano/genética , Células HEK293 , Humanos , Sequências Repetidas Invertidas , Staphylococcus/química , Staphylococcus aureus/enzimologia
18.
Nat Microbiol ; 5(4): 620-629, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32218510

RESUMO

CRISPR-Cas adaptive immune systems protect bacteria and archaea against their invading genetic parasites, including bacteriophages/viruses and plasmids. In response to this immunity, many phages have anti-CRISPR (Acr) proteins that inhibit CRISPR-Cas targeting. To date, anti-CRISPR genes have primarily been discovered in phage or prophage genomes. Here, we uncovered acr loci on plasmids and other conjugative elements present in Firmicutes using the Listeria acrIIA1 gene as a marker. The four identified genes, found in Listeria, Enterococcus, Streptococcus and Staphylococcus genomes, can inhibit type II-A SpyCas9 or SauCas9, and are thus named acrIIA16-19. In Enterococcus faecalis, conjugation of a Cas9-targeted plasmid was enhanced by anti-CRISPRs derived from Enterococcus conjugative elements, highlighting a role for Acrs in the dissemination of plasmids. Reciprocal co-immunoprecipitation showed that each Acr protein interacts with Cas9, and Cas9-Acr complexes were unable to cleave DNA. Northern blotting suggests that these anti-CRISPRs manipulate single guide RNA length, loading or stability. Mirroring their activity in bacteria, AcrIIA16 and AcrIIA17 provide robust and highly potent broad-spectrum inhibition of distinct Cas9 proteins in human cells (for example, SpyCas9, SauCas9, SthCas9, NmeCas9 and CjeCas9). This work presents a focused analysis of non-phage Acr proteins, demonstrating a role in horizontal gene transfer bolstered by broad-spectrum CRISPR-Cas9 inhibition.


Assuntos
Proteína 9 Associada à CRISPR/antagonistas & inibidores , Sistemas CRISPR-Cas , Transferência Genética Horizontal , Plasmídeos/metabolismo , RNA Guia/antagonistas & inibidores , Proteína 9 Associada à CRISPR/genética , Proteína 9 Associada à CRISPR/metabolismo , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Conjugação Genética , DNA/antagonistas & inibidores , DNA/genética , DNA/metabolismo , Enterococcus/genética , Enterococcus/virologia , Células HEK293 , Humanos , Listeria/genética , Listeria/virologia , Plasmídeos/química , Ligação Proteica , RNA Guia/genética , RNA Guia/metabolismo , Staphylococcus/genética , Staphylococcus/virologia , Streptococcus/genética , Streptococcus/virologia
19.
Food Microbiol ; 89: 103434, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32138992

RESUMO

Acidification level and temperature modulate the beneficial consortia of lactic acid bacteria (LAB) and coagulase-negative staphylococci (CNS) during meat fermentation. Less is known about the impact of other factors, such as raw meat quality and salting. These could for instance affect the growth of the pathogen Staphylococcus aureus or of Enterobacterales species, potentially indicative of poor fermentation practice. Therefore, pork batters from either normal or borderline quality (dark-firm-dry, DFD) were compared at various salt concentrations (0-4%) in meat fermentation models. Microbial ecology of the samples was investigated with culture-dependent techniques and (GTG)5-PCR fingerprinting of genomic DNA. Whilst Lactobacillus sakei governed the fermentation of normal meat, Lactobacillus curvatus was more prominent in the fermentation of the DFD meat variant. CNS were favoured during fermentation at rising salt concentrations without much effects on species diversity, consisting mostly of Staphylococcus equorum, Staphylococcus saprophyticus, and Staphylococcus xylosus. During fermentation of DFD meat, S. saprophyticus was less manifest than during that of normal meat. Enterobacterales mainly emerged in DFD meat during fermentation at low salt concentrations. The salt hurdle was insufficient to prevent Enterobacterales when acidification and initial pH were favourable for their growth.


Assuntos
Microbiologia de Alimentos , Lactobacillus/genética , Produtos da Carne/microbiologia , Carne de Porco/microbiologia , Staphylococcus/genética , Animais , Fermentação , Cloreto de Sódio na Dieta , Suínos
20.
PLoS One ; 15(2): e0228199, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32040520

RESUMO

In the present study, we identified salt stress tolerant genes from the marine bacterium Staphylococcus sp. strain P-TSB-70 through transcriptome sequencing. In favour of whole-genome transcriptome profiling of Staphylococcus sp. strain P-TSB-70 (GenBank Accn. No. KP117091) which tolerated upto 20% NaCl stress, the strain was cultured in the laboratory condition with 20% NaCl stress. Transcriptome analyses were performed by SOLiD4.0 sequencing technology from which 10280 and 9612 transcripts for control and treated, respectively, were obtained. The coverage per base (CPB) statistics were analyzed for both the samples. Gene ontology (GO) analysis has been categorized at varied graph levels based on three primary ontology studies viz. cellular components, biological processes, and molecular functions. The KEGG analysis of the assembled transcripts using KAAS showed presumed components of metabolic pathways which perhaps implicated in diverse metabolic pathways responsible for salt tolerance viz. glycolysis/gluconeogenesis, oxidative phosphorylation, glutathione metabolism, etc. further involving in salt tolerance. Overall, 90 salt stress tolerant genes were identified as of 186 salt-related transcripts. Several genes have been found executing normally in the TCA cycle pathway, integral membrane proteins, generation of the osmoprotectants, enzymatic pathway associated with salt tolerance. Recognized genes fit diverse groups of salt stress genes viz. abc transporter, betaine, sodium antiporter, sodium symporter, trehalose, ectoine, and choline, that belong to different families of genes involved in the pathway of salt stress. The control sample of the bacterium showed elevated high proportion of transcript contigs (29%) while upto 20% salt stress treated sample of the bacterium showed a higher percentage of transcript contigs (31.28%). A total of 1,288 and 1,133 transcript contigs were measured entirely as novel transcript contigs in both control and treated samples, respectively. The structure and function of 10 significant salt stress tolerant genes of Staphylococcus sp. have been analyzed in this study. The information acquired in the present study possibly used to recognize and clone the salt stress tolerant genes and support in developing the salt stress-tolerant plant varieties to expand the agricultural productivity in the saline system.


Assuntos
Tolerância ao Sal/genética , Água do Mar/microbiologia , Staphylococcus/genética , Transcriptoma , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Mapeamento de Sequências Contíguas , Perfilação da Expressão Gênica , Genoma Bacteriano , Índia , Redes e Vias Metabólicas/genética , Estrutura Terciária de Proteína , RNA Bacteriano/química , RNA Bacteriano/metabolismo , Staphylococcus/isolamento & purificação
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