Studies on metal resistant bacteria in cochin estuary and its response towards antibiotics and silver nanoparticles

The Cochin estuary (CE), which is one of the largest wetland ecosystems, extends from Thanneermukkam bund in the south to Azhikode in the north. It functions as an effluent repository for more than 240 industries, the characteristics of which includes fertilizer, pesticide, radioactive mineral processing, chemical and allied industries, petroleum refining and heavy metal processing industries (Thyagarajan, 2004). Studies in the CE have been mostly on the spatial and temporal variations in the physical, chemical and biological characteristics of the estuary (Balachandran et al., 2006; Madhu et al., 2007; Menon et al., 2000; Qasim 2003;Qasim and Gopinathan 1969) . Although several monitoring programs have been initiated in the CE to understand the level of heavy metal pollution, these were restricted to trace metals distribution (Balachandran et al., 2005) or the influence of anthropogenic inputs on the benthos and phytoplankton (Madhu et al., 2007;Jayaraj, 2006). Recently, few studies were carried out on microbial ecology in the CE(Thottathil et al 2008a and b;Parvathi et al., 2009and 2011; Thomas et al., 2006;Chandran and Hatha, 2003). However, studies on metal - microbe interaction are hitherto not undertaken in this estuary. Hence, a study was undertaken at 3 sites with different level of heavy metal concentration tounderstand the abundance, diversity and mechanisms of resistance in metal resistant bacteria and its impact on the nutrient regeneration. The present work has also focused on the response of heavy metal resistant bacteria towards antibacterial agent’s antibiotics and silver nanoparticles

The hyperaccumulator Sedum plumbizincicola harbors metal-resistant endophytic bacteria that improve its phytoextraction capacity in multi-metal contaminated soil

Endophyte-assisted phytoremediation has recently been suggested as a successful approach for ecological restoration of metal contaminated soils, however little information is available on the influence of endophytic bacteria on the phytoextraction capacity of metal hyperaccumulating plants in multi-metal polluted soils. The aims of our study were to isolate and characterize metal-resistant and 1-aminocyclopropane-1-carboxylate (ACC) utilizing endophytic bacteria from tissues of the newly discovered Zn/Cd hyperaccumulator Sedum plumbizincicola and to examine if these endophytic bacterial strains could improve the efficiency of phytoextraction of multi-metal contaminated soils. Among a collection of 42 metal resistant bacterial strains isolated from the tissues of S. plumbizincicola grown on Pb/Zn mine tailings, five plant growth promoting endophytic bacterial strains (PGPE) were selected due to their ability to promote plant growth and to utilize ACC as the sole nitrogen source. The five isolates were identified as Bacillus pumilus E2S2, Bacillus sp. E1S2, Bacillus sp. E4S1, Achromobacter sp. E4L5 and Stenotrophomonas sp. E1L and subsequent testing revealed that they all exhibited traits associated with plant growth promotion, such as production of indole-3-acetic acid and siderophores and solubilization of phosphorus. These five strains showed high resistance to heavy metals (Cd, Zn and Pb) and various antibiotics. Further, inoculation of these ACC utilizing strains significantly increased the concentrations of water extractable Cd and Zn in soil. Moreover, a pot experiment was conducted to elucidate the effects of inoculating metal-resistant ACC utilizing strains on the growth of S. plumbizincicola and its uptake of Cd, Zn and Pb in multi-metal contaminated soils. Out of the five strains, B. pumilus E2S2 significantly increased root (146%) and shoot (17%) length, fresh (37%) and dry biomass (32%) of S. plumbizincicola as well as plant Cd uptake (43%), whereas Bacillus sp. E1S2 significantly enhanced the accumulation of Zn (18%) in plants compared with non-inoculated controls. The inoculated strains also showed high levels of colonization in rhizosphere and plant tissues. Results demonstrate the potential to improve phytoextraction of soils contaminated with multiple heavy metals by inoculating metal hyperaccumulating plants with their own selected functional endophytic bacterial strains.

Heavy metal tolerance patterns of total heterotrophic bacteria isolated from the soils of Mahatma Gandhi University campus, Kottayam, Kerala

Industrialization of our society has led to an increased production and discharge of both xenobiotic and natural chemical substances. Many of these chemicals will end up in the soil. Pollution of soils with heavy metals is becoming one of the most severe ecological and human health hazards. Elevated levels of heavy metals decrease soil microbial activity and bacteria need to develop different mechanisms to confer resistances to these heavy metals. Bacteria develop heavy-metal resistance mostly for their survivals, especially a significant portion of the resistant phenomena was found in the environmental strains. Therefore, in the present work, we check the multiple metal tolerance patterns of bacterial strains isolated from the soils of MG University campus, Kottayam. A total of 46 bacterial strains were isolated from different locations of the campus and tested for their resistant to 5 common metals in use (lead, zinc, copper, cadmium and nickel) by agar dilution method. The results of the present work revealed that there was a spatial variation of bacterial metal resistance in the soils of MG University campus, this may be due to the difference in metal contamination in different sampling location. All of the isolates showed resistance to one or more heavy metals selected. Tolerance to lead was relatively high followed by zinc, nickel, copper and cadmium. About 33% of the isolates showed very high tolerance (>4000μg/ml) to lead. Tolerance to cadmium (65%) was rather low (<100 μg/ml). Resistance to zinc was in between 100μg/ml - 1000μg/ml and the majority of them shows resistance in between 200μg/ml - 500μg/ml. Nickel resistance was in between 100μg/ml - 1000μg/ml and a good number of them shows resistance in between 300μg/ml - 400μg/ml. Resistance to copper was in between <100μg/ml - 500μg/ml and most of them showed resistance in between 300μg/ml - 400μg/ml. From the results of this study, it was concluded that heavy metal-resistant bacteria are widely distributed in the soils of MG university campus and the tolerance of heavy metals varied among bacteria and between locations

Design of Antibacterial Prochelators to Target Drug-Resistant Bacteria

Transition metals such as iron and copper are valued in biology for their redox activities because they are able to access various oxidation states. However, these transition metals are also implicated in a number of human disease states and play a role in bacterial infections. The ability to manipulate and monitor metal ions has vast implications on the fields of biology and human health. As such, the research described here covers two related goals: to manipulate metals in specific biological circumstances and to visualize this disturbance in cellular metal homeostasis.

Antibiotic resistance necessitates the development of drugs that exploit new mechanisms of action such as the disruption of metal homeostasis. In order to manipulate metals at the site of bacterial infection, two prochelators were developed around a β-lactam core such that the active chelator is released in the presence of bacteria that produce the resistance-causing β-lactamase enzyme. Both prochelators display enhanced activity toward resistant bacteria compared to clinical antibiotics.

Fluorescent sensors are a powerful tool for detecting small concentrations of biological analytes. Two analogs of a ratiometric fluorescent sensor were designed and synthesized to monitor cellular concentrations of copper and iron. These sensors were found to operate as designed in vitro; however the fluorescence intensity necessary for quantification of cellular metal pools has not yet been achieved.

Antibacterial activity of Ionic Liquids based on ampicillin against resistant bacteria

Antibacterial activity of novel Active Pharmaceutical Ingredient Ionic Liquids (API-ILs) based on ampicillin anion [Amp] have been evaluated. They showed growth inhibition and bactericidal properties on some sensitive bacteria and especially some Gram-negative resistant bacteria when compared to the [Na][Amp] and the initial bromide and chloride salts. For these studies were analysed the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBIC) against sensitive Gram-negative bacteria Escherichia coli ATCC 25922 and Klebsiella pneumonia (clinically isolated), as well as sensitive Gram positive S. Aureus ATCC 25923, Staphylococcus epidermidis and Enterococcus faecalis and completed using clinically isolated resistent strains: E. coli TEM CTX M9, E. coli CTX M2 and E. coli AmpC Mox. From the obtained MIC values of studied APIs-ILs and standard [Na][Amp] were derived RDIC values (relative decrease of inhibitory concentration). High RDIC values of [C16Pyr][Amp] especially against two resistant Gram-negative strains E. coli TEM CTX M9 (RDIC>1000) and E. coli CTX M2 (RDIC>100) point clearly to a potential promising role of APIs-ILs as antimicrobial drugs especially against resistant bacterial strains.

The inhibitory activity of synthetic compounds and ions against transporters of multi-drug resistant bacteria

RESUMO: Sessenta e três derivados de hidantoína foram utilizados para avaliar possíveis efeitos de modulação na actividade das bombas de efluxo (BE) na Salmonella NCTC 13349 utilizando um método fluorimétrico semi-automático. Nenhum dos compostos apresentaram actividade anti-bacteriana até concentrações de 240 mg/L. Entre todos os compostos, SZ-7 demonstrou possuir propriedades de modulação de effluxo na presença de glucose. Para testar esta actividade, estirpes de Salmonella resistentes à ciprofloxacina, induzidas a elevados níveis de resistência com sobre-expressão de BE, foram expostas ao SZ-7. Este derivado afectou a susceptibilidade das estirpes à ciprofloxacina. Uma vez que os 63 compostos estudados apresentaram pouco efeito inibitório /cumulativo, apesar de serem conhecidos pelos seus efeitos moduladores de BE-dependentes de iões em eucariotas, foi questionado o papel dos iões na regulação de BE bacterianas, que poderão influenciar a eficácia de novos compostos. Para este estudo, utilizamos a Escherichia coli AG100 como modelo, devido ao extenso conhecimento no que respeita a estrutura e actividade das BE. Devido à importância de iões de cálcio (Ca2+) nos canais de transporte membranar e na actividade de ATPases, a sua actividade na modulação do efluxo foi investigada. De resultados anteriormente obtidos concluiu-se que a pH 5 o efluxo é independente de energia metabólica; contudo, a pH 8 é absolutamente dependente, sendo que o Ca2+ é indispensável para manter a actividade das ATPases bacterianas. A acumulação/effluxo de EtBr pela E. coli AG100 foi determinada na presença/ausência de Ca2+, clorpromazina (inibidor de ligação de Ca2+ a proteínas), e ácido etilenodiamino tetra-acético (quelante de Ca2+). Acumulação/effluxo aumentou a pH 8, contudo o Ca2+ reverte estes efeitos evidenciando a sua importância no funcionamento das BE bacterianas. Em resumo este trabalho colocou em evidência que muitos aspectos bioquímicos e bioenergéticos devem ser tomados em consideração no estudo da resistência bacteriana mediada por BE.------- ABSTRACT: Sixty-three hydantoin derivatives were evaluated for their modulating effects on efflux pump (EP) activity of Salmonella NCTC 13349 utilizing a semi-automatic fluorometric method. None of the compounds presented antibacterial activities at concentrations as high as 240 mg/L. Among all compounds, SZ-7 showed possible efflux modulating activity in the presence of glucose, indicative of a potential EP inhibitor. To verify its potential effects, ciprofloxacin-resistant Salmonella strains, induced to high level resistance with over-expressing EPs, were exposed to SZ-7. This derivative affected the susceptibility of the ciprofloxacin-resistant strains. Since the 63 compounds studied had very low inhibitory/accumulative effects, even though their known for being efficient in modulating ion-driven eukaryotic EPs, we questioned whether ions had a leading role in regulating bacterial EPs, influencing the effectiveness of new compounds. For this study we used Escherichia coli AG100 as a model, due to the extensive knowledge on its EPs structure and activity. Owing the importance of calcium ions (Ca2+) for membrane transport channels and activity of ATPases, the role of Ca2+ was investigated. From previous results we concluded that at pH 5 efflux is independent of metabolic energy; however, at pH 8 it is entirely dependent of metabolic energy and the Ca2+ ions are essential to maintain the activity of bacterial ATPases. Accumulation and efflux of ethidium bromide (EtBr) by E. coli AG100 was determined in the presence and absence of Ca2+, chlorpromazine (inhibitor of Ca2+-binding to proteins), and ethylenediaminetetraacetic acid (Ca2+ chelator). Accumulation of EtBr increased at pH 8; however Ca2+ reversed these effects providing information as to the importance of this ion in the regulation of bacterial EP systems. Overall this work puts in evidence that many biochemical and bioenergetic aspects related to the strains physiology need to be taken into consideration in bacterial drug resistance mediated by EPs.

New methods for detecting antibiotic resistant bacteria and preventing their spread

Methicillin resistant Staphylococcus aureus (MRSA) bacteria have emerged in the early 1980's in numerous health care institutions around the world. The main transmission mechanism within hospitals and healthcare facilities is through the hands of health care workers. Resistant to several antibiotics, the MRSA is one of the most feared pathogens in the hospital setting since it is very difficult to eradicate with the standard treatments. There are still a limited number of anti-MRSA antibiotics but the first cases of resistance to these compounds have already been reported and their frequency is likely to increase in the coming years. Every year, the MRSA infections result in major human and financial costs, due to the high associated mortality and expenses related to the required care. Measures towards a faster detection of resistant bacteria and establishment of appropriate antibiotic treatment parameters are fundamental. Also as part as infection prevention, diminution of bacteria present on the commonly touched surfaces could also limit the spread and selection of antibiotic resistant bacteria. During my thesis, projects were developed around MRSA and antibiotic resistance investigation using innovative technologies. The thesis was subdivided in three main parts with the use of atomic force microscopy AFM for antibiotic resistance detection in part 1, the importance of the bacterial inoculum size in the selection of antibiotic resistance in part 2 and the testing of antimicrobial surfaces creating by sputtering copper onto polyester in part 3. In part 1 the AFM was used two different ways, first for the measurement of stiffness (elasticity) of bacteria and second as a nanosensor for antibiotic susceptibility testing. The stiffness of MRSA with different susceptibility profiles to vancomycin was investigated using the stiffness tomography mode of the AFM and results have demonstrated and increased stiffness in the vancomycin resistant strains that also paralleled with increased thickness of the bacterial cell wall. Parts of the AFM were also used to build a new antibiotic susceptibility-testing device. This nano sensor was able to measure vibrations emitted from living bacteria that ceased definitively upon antibiotic exposure to which they were susceptible but restarted after antibiotic removal to which they were resistant, allowing in a matter of minute the assessment of antibiotic susceptibility determination. In part 2 the inoculum effect (IE) of vancomycin, daptomycin and linezolid and its importance in antibiotic resistance selection was investigated with MRSA during a 15 days of cycling experiment. Results indicated that a high bacterial inoculum and a prolonged antibiotic exposure were two key factors in the in vitro antibiotic resistance selection in MRSA and should be taken into consideration when choosing the drug treatment. Finally in part 3 bactericidal textile surfaces were investigated against MRSA. Polyesters coated after 160 seconds of copper sputtering have demonstrated a high bactericidal activity reducing the bacterial load of at least 3 logio after one hour of contact. -- Au cours des dernières décennies, des bactéries multirésistantes aux antibiotiques (BMR) ont émergé dans les hôpitaux du monde entier. Depuis lors, le nombre de BMR et la prévalence des infections liées aux soins (IAS) continuent de croître et sont associés à une augmentation des taux de morbidité et de mortalité ainsi qu'à des coûts élevés. De plus, le nombre de résistance à différentes classes d'antibiotiques a également augmenté parmi les BMR, limitant ainsi les options thérapeutiques disponibles lorsqu'elles ont liées a des infections. Des mesures visant une détection plus rapide des bactéries résistantes ainsi que l'établissement des paramètres de traitement antibiotiques adéquats sont primordiales lors d'infections déjà présentes. Dans une optique de prévention, la diminution des bactéries présentes sur les surfaces communément touchées pourrait aussi freiner la dissémination et l'évolution des bactéries résistantes. Durant ma thèse, différents projets incluant des nouvelles technologies et évoluant autour de la résistance antibiotique ont été traités. Des nouvelles technologies telles que le microscope à force atomique (AFM) et la pulvérisation cathodique de cuivre (PCC) ont été utilisées, et le Staphylococcus aureus résistant à la méticilline (SARM) a été la principale BMR étudiée. Deux grandes lignes de recherche ont été développées; la première visant à détecter la résistance antibiotique plus rapidement avec l'AFM et la seconde visant à prévenir la dissémination des BMR avec des surfaces crées grâce à la PCC. L'AFM a tout d'abord été utilisé en tant que microscope à sonde locale afin d'investiguer la résistance à la vancomycine chez les SARMs. Les résultats ont démontré que la rigidité de la paroi augmentait avec la résistance à la vancomycine et que celle-ci corrélait aussi avec une augmentation de l'épaisseur des parois, vérifiée grâce à la microscopie électronique. Des parties d'un AFM ont été ensuite utilisées afin de créer un nouveau dispositif de test de sensibilité aux antibiotiques, un nanocapteur. Ce nanocapteur mesure des vibrations produites par les bactéries vivantes. Après l'ajout d'antibiotique, les vibrations cessent définitivement chez les bactéries sensibles à l'antibiotique. En revanche pour les bactéries résistantes, les vibrations reprennent après le retrait de l'antibiotique dans le milieu permettant ainsi, en l'espace de minutes de détecter la sensibilité de la bactérie à un antibiotique. La PCC a été utilisée afin de créer des surfaces bactéricides pour la prévention de la viabilité des BMR sur des surfaces inertes. Des polyesters finement recouverts de cuivre (Cu), connu pour ses propriétés bactéricides, ont été produits et testés contre des SARMs. Une méthode de détection de viabilité des bactéries sur ces surfaces a été mise au point, et les polyesters obtenus après 160 secondes de pulvérisation au Cu ont démontré une excellente activité bactéricide, diminuant la charge bactérienne d'au moins 3 logio après une heure de contact. En conclusion, l'utilisation de nouvelles technologies nous a permis d'évoluer vers de méthodes de détection de la résistance antibiotique plus rapides ainsi que vers le développement d'un nouveau type de surface bactéricide, dans le but d'améliorer le diagnostic et la gestion des BMR.

Extended spectrum beta lactamase (ESBL) resistant bacteria - including accessible formats

Information for patients and visitors on extended spectrum beta lactamase (ESBL) resistant bacteria and how to help prevent the spread of infection.Accessible formatsThe below document is available as a pdf and in accessible formats. Accessible formats are alternatives to printed information, used by people who are blind or visually impaired. These accessible formats include HTML, audio and braille. �For audio and HTML copies please click on the links below. For braille copies please contact Caroline McGeary on 0300 555 0114.

Marine Pseudomonas putida: a potential source of antimicrobial substances against antibiotic-resistant bacteria

Bacteria isolated from marine sponges found off the coast of Rio de Janeiro, Brazil, were screened for the production of antimicrobial substances. We report a new Pseudomonas putida strain (designated P. putida Mm3) isolated from the sponge Mycale microsigmatosa that produces a powerful antimicrobial substance active against multidrug-resistant bacteria. P. putida Mm3 was identified on the basis of 16S rRNA gene sequencing and phenotypic tests. Molecular typing for Mm3 was performed by RAPD-PCR and comparison of the results to other Pseudomonas strains. Our results contribute to the search for new antimicrobial agents, an important strategy for developing alternative therapies to treat infections caused by multidrug-resistant bacteria.

Antibiotic resistant bacteria/genes dissemination in lacustrine sediments highly increased following cultural eutrophication of Lake Geneva (Switzerland).

This study investigates faecal indicator bacteria (FIB), multiple antibiotic resistant (MAR), and antibiotic resistance genes (ARGs), of sediment profiles from different parts of Lake Geneva (Switzerland) over the last decades. MARs consist to expose culturable Escherichia coli (EC) and Enterococcus (ENT) to mixed five antibiotics including Ampicillin, Tetracycline, Amoxicillin, Chloramphenicol and Erythromycin. Culture-independent is performed to assess the distribution of ARGs responsible for, β-lactams (blaTEM; Amoxicillin/Ampicillin), Streptomycin/Spectinomycin (aadA), Tetracycline (tet) Chloramphenicol (cmlA) and Vancomycin (van). Bacterial cultures reveal that in the sediments deposited following eutrophication of Lake Geneva in the 1970s, the percentage of MARs to five antibiotics varied from 0.12% to 4.6% and 0.016% to 11.6% of total culturable EC and ENT, respectively. In these organic-rich bacteria-contaminated sediments, the blaTEM resistant of FIB varied from 22% to 48% and 16% to 37% for EC and ENT respectively, whereas the positive PCR assays responsible for tested ARGs were observed for EC, ENT, and total DNA from all samples. The aadA resistance gene was amplified for all the sediment samples, including those not influenced by WWTP effluent water. Our results demonstrate that bacteria MARs and ARGs highly increased in the sediments contaminated with WWTP effluent following the cultural eutrophication of Lake Geneva. Hence, the human-induced changing limnological conditions highly enhanced the sediment microbial activity, and therein the spreading of antibiotic resistant bacteria and genes in this aquatic environment used to supply drinking water in a highly populated area. Furthermore, the presence of the antibiotic resistance gene aadA in all the studied samples points out a regional dissemination of this emerging contaminant in freshwater sediments since at least the late nineteenth century.

Targeted therapy against multi-resistant bacteria in leukemic and hematopoietic stem cell transplant recipients: guidelines of the 4th European Conference on Infections in Leukemia (ECIL-4, 2011).

The detection of multi-resistant bacterial pathogens, particularly those to carbapenemases, in leukemic and stem cell transplant patients forces the use of old or non-conventional agents as the only remaining treatment options. These include colistin/polymyxin B, tigecycline, fosfomycin and various anti-gram-positive agents. Data on the use of these agents in leukemic patients are scanty, with only linezolid subjected to formal trials. The Expert Group of the 4(th) European Conference on Infections in Leukemia has developed guidelines for their use in these patient populations. Targeted therapy should be based on (i) in vitro susceptibility data, (ii) knowledge of the best treatment option against the particular species or phenotype of bacteria, (iii) pharmacokinetic/pharmacodynamic data, and (iv) careful assessment of the risk-benefit balance. For infections due to resistant Gram-negative bacteria, these agents should be preferably used in combination with other agents that remain active in vitro, because of suboptimal efficacy (e.g., tigecycline) and the risk of emergent resistance (e.g., fosfomycin). The paucity of new antibacterial drugs in the near future should lead us to limit the use of these drugs to situations where no alternative exists.

Preventive and therapeutic strategies in critically ill patients with highly resistant bacteria.

The antibiotic pipeline continues to diminish and the majority of the public remains unaware of this critical situation. The cause of the decline of antibiotic development is multifactorial and currently most ICUs are confronted with the challenge of multidrug-resistant organisms. Antimicrobial multidrug resistance is expanding all over the world, with extreme and pandrug resistance being increasingly encountered, especially in healthcare-associated infections in large highly specialized hospitals. Antibiotic stewardship for critically ill patients translated into the implementation of specific guidelines, largely promoted by the Surviving Sepsis Campaign, targeted at education to optimize choice, dosage, and duration of antibiotics in order to improve outcomes and reduce the development of resistance. Inappropriate antimicrobial therapy, meaning the selection of an antibiotic to which the causative pathogen is resistant, is a consistent predictor of poor outcomes in septic patients. Therefore, pharmacokinetically/pharmacodynamically optimized dosing regimens should be given to all patients empirically and, once the pathogen and susceptibility are known, local stewardship practices may be employed on the basis of clinical response to redefine an appropriate regimen for the patient. This review will focus on the most severely ill patients, for whom substantial progress in organ support along with diagnostic and therapeutic strategies markedly increased the risk of nosocomial infections.

Bottled mineral water as a potential source of antibiotic resistant bacteria

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

In vitro antibacterial and chemical properties of essential oils including native plants from Brazil against pathogenic and resistant bacteria

The antimicrobials products from plants have increased in importance due to the therapeutic potential in the treatment of infectious diseases. Therefore, we aimed to examine the chemical characterisation (GC-MS) of essential oils (EO) from seven plants and measure antibacterial activities against bacterial strains isolated from clinical human specimens (methicillin-resistant Staphylococcus aureus (MRSA) and sensitive (MSSA), Escherichia coli, Pseudomonas aeruginosa, Salmonella Typhimurium) and foods (Salmonella Enteritidis). Assays were performed using the minimal inhibitory concentration (MIC and MIC90%) (mg/mL) by agar dilution and time kill curve methods (log CFU/mL) to aiming synergism between EO. EO chemical analysis showed a predominance of terpenes and its derivatives. The highest antibacterial activities were with Cinnamomun zeylanicum (0.25 mg/mL on almost bacteria tested) and Caryophyllus aronzaticus EO (2.40 mg/mL on Salmonella Enteritidis), and the lowest activity was with Eugenia uniflora (from 50.80 mg/mL against MSSA to 92.40 mg/mL against both Salmonella sources and P aeruginosa) EO. The time kill curve assays revealed the occurrence of bactericide synergism in combinations of C. aromaticus and C. zeylanicum with Rosmarinus. officinalis. Thus, the antibacterial activities of the EO were large and this can also be explained by complex chemical composition of the oils tested in this study and the synergistic effect of these EO, yet requires further investigation because these interactions between the various chemical compounds can increase or reduce (antagonism effect) the inhibitory effect of essential oils against bacterial strains.