Hemolysin Production Research Articles

Optimization of hemolysin formation in Alcaligenes species isolated from abattoir wastewater samples in Akure, Ondo State, Nigeria

Hemolysin is significantly toxic and it is used as a molecular marker for pathogenicity. This study evaluates the conditions for optimal hemolysin production by Alcaligenes faecalis strains isolated from a city abattoir wastewater. The parameters investigated for hemolysin formation were size of the inoculum, initial pH of production medium, bacterial incubation temperature, agitation speed and growth media. Thereafter, the effect by various parameter on the hemolytic activity for the formation of hemolysin were assessed. The genus Alcaligenes was assigned to the test organisms after analyzing their 16S rRNA gene sequence with accession numbers: MF498824, MF498825 and MF498827. Optimum conditions for hemolysin formation in Alcaligenes faecalis strain OS42 were inoculum size of 0.5% (v/v), pH 9, 20 oC, 0 rpm and brain heart infusion broth. Hemolytic activities, 77% and 79% were achieved at 20 h for strains OS42 and OS61, respectively. Cholesterol and ethylenediaminetetraacetic acid did not affect hemolysin formation. This work revealed that the hemolysin formation in Alcaligenes strains was sourced from abattoir wastewater effluent and the effluent was contaminated with pathogenic Alcaligenes strains which is a public health hazard due to their prospective infection to human and animal.

THE RELATIONSHIP BETWEEN PRODUCTION OF PROTEIN A, CAPSULE STRUCTURE AND SOME ENZYMES OF STAPHYLOCOCCUS AUREUS ISOLATED FROM INFECTIONS OF HUMAN AND ANIMALS

Thirty-five isolates of Staphylococcus were isolated out of 100 clinical specimens were collected from different infections of human and animals. These isolates were identified as Staphylococcus aureus, 15 of them isolated from human, 15 from cows and only 5 isolates from poultry. Twenty-five (71.4%) isolates of different sources revealed positive results for the detection of protein A, capsule formation and enzyme production of phosphatase, DNase and haemolysin. Seven (20 %) isolates showed negative results for both protein A and capsule as well as for the production of enzymes while only three (8.5%) isolates were not capsulated but able to produce protein A and the enzymes. During the detection of protein A, capsule structure and production of some enzymes, we found that the best value of pH used was the range of 7-7.5. The present study indicated that the presence of protein A and capsule structure is highly associated with the virulence of isolates, and loss of protein A doesn't necessary mean loss of capsule as well. The ability of the isolates to produce their essential enzymes to induct the infection is highly correlated with the ability to produce protein A, without considering the source of the isolates.

Inhibition of Quorum Sensing and Biofilm Formation of Esculetin on Aeromonas Hydrophila.

Quorum sensing (QS) and biofilm formation inhibition activity of esculetin on Aeromonas hydrophila SHAe 115 were evaluated. Exposure to esculetin at 25, 50, and 100μg/ml significantly inhibited the production of protease and hemolysin, the formation of biofilms and attenuated the swarming motility of A. hydrophila SHAe 115. Biofilm forming inhibition was also observed through confocal laser scanning microscopy and scanning electron microscope. Quantitative real-time PCR analysis indicated that genes positively related to QS and biofilm formation were downregulated to varying degrees, while gene (litR) negatively related to biofilm formation was significantly upregulated. The phenotypic results were in good agreement with gene expression levels. These results indicated that esculetin would be a potential QS inhibitor for A. hydrophila.

Molecular Epidemiology, Antifungal Susceptibility, and Virulence Evaluation of Candida Isolates Causing Invasive Infection in a Tertiary Care Teaching Hospital.

BackgroundThe incidence of invasive candidiasis is increasing worldwide. However, the epidemiology, antifungal susceptibility, and virulence of Candida spp. in most hospitals remain unclear. This study aimed to evaluate invasive candidiasis in a tertiary care hospital in Nanchang City, China.MethodsMALDI-TOF MS and 18S rDNA ITS sequencing were used to identify Candida strains. Randomly amplified polymorphic DNA analysis was used for molecular typing; biofilm production, caseinase, and hemolysin activities were used to evaluate virulence. The Sensititre™ YeastOne YO10 panel was used to examine antifungal susceptibility. Mutations in ERG11 and the hotspot regions of FKS1 of drug-resistant strains were sequenced to evaluate the possible mechanisms of antifungal resistance.ResultsWe obtained 110 Candida strains, which included 40 Candida albicans (36.36%), 37 C. parapsilosis (33.64%), 21 C. tropicalis (19.09%), 9 C. glabrata (8.18%), 2 C. rugose (1.82%), and 1 C. haemulonii (0.91%) isolates. At a limiting point of 0.80, C. albicans isolates could be grouped into five clusters, C. parapsilosis and C. tropicalis isolates into seven clusters, and C. glabrata isolates into only one cluster comprising six strains by RAPD typing. Antifungal susceptibility testing revealed that the isolates showed the greatest overall resistance against fluconazole (6.36%), followed by voriconazole (4.55%). All C. albicans and C. parapsilosis isolates exhibited 100% susceptibility to echinocandins (i.e., anidulafungin, caspofungin, and micafungin), whereas one C. glabrata strain was resistant to echinocandins. The most common amino acid substitutions noted in our study was 132aa (Y132H, Y132F) in the azole-resistant strains. No missense mutation was identified in the hotpot regions of FKS1. Comparison of the selected virulence factors detectable in a laboratory environment, such as biofilm, caseinase, and hemolysin production, revealed that most Candida isolates were caseinase and hemolysin producers with a strong activity (Pz < 0.69). Furthermore, C. parapsilosis had greater total biofilm biomass (average Abs620 = 0.712) than C. albicans (average Abs620 = 0.214, p < 0.01) or C. tropicalis (average Abs620 = 0.450, p < 0.05), although all C. glabrata strains were either low- or no-biofilm producers. The virulence level of the isolates from different specimen sources or clusters showed no obvious correlation. Interesting, 75% of the C. albicans from cluster F demonstrated azole resistance, whereas two azole-resistant C. tropicalis strains belonged to the cluster Y.ConclusionThis study provides vital information regarding the epidemiology, pathogenicity, and antifungal susceptibility of Candida spp. in patients admitted to Nanchang City Hospital.

Probiotic potential and safety assessment of bacteriocinogenic Enterococcus faecium strains with antibacterial activity against Listeria and vancomycin-resistant enterococci

Enterococcus spp., known for their wide ecological distribution, have been associated with various fermented food products of plant and animal origin. The strains used in this study, bacteriocinogenic Enterococcus faecium previously isolated from artisanal soybean paste, have shown strong activity against Listeria spp. and vancomycin-resistant enterococci. Although their antimicrobial activity is considered beneficial, the potential application of enterococci is still under debate due to concerns about their safety for human and other animal consumption. Therefore, this study not only focuses on the screening of potential virulence factors, but also the auxiliary beneficial properties of the strains Ent. faecium ST651ea, ST7119ea, and ST7319ea. Phenotypic screening for gelatinase, hemolysin, and biogenic amine production showed that the strains were all safe. Furthermore, the antibiogram profiling showed that all the strains were susceptible to the panel of antibiotics used in the assessment except for erythromycin. Yet, Ent. faecium ST7319ea was found to carry some of the virulence genes used in the molecular screening for safety including hyl, esp, and IS16. The probiotic potential and other beneficial properties of the strains were also studied, demonstrating high aggregation and co-aggregation levels compared to previously characterized strains, in addition to high survivability under simulated gastrointestinal conditions, and production of numerous desirable enzymes as evaluated by APIZym, indicating diverse possible biotechnological applications of these strains. Additionally, the strains were found to carry genes coding for γ-aminobutyric acid (GABA) production, an auxiliary characteristic for their probiotic potential. Although these tests showed relatively favorable characteristics, it should be considered that these assays were carried out in vitro and should therefore also be assessed under in vivo conditions.

Virulence Factors and Azole-Resistant Mechanism of Candida Tropicalis Isolated from Candidemia.

Limited knowledge exists on the virulence factors of Candida tropicalis and the mechanisms of azole resistance that lead to an intensified pathogenicity and treatment failure. We aimed to evaluate the virulence factors and molecular mechanisms of azole resistance among C. tropicalis isolated from patients with candidemia. Several virulence factors, including extracellular enzymatic activities, cell surface hydrophobicity (CSH), and biofilm formation, were evaluated. Antifungal susceptibility pattern and expression level of ERG11, UPC2, MDR1, and CDR1 genes of eight (4 fluconazole resistance and 4 fluconazole susceptible) clinical C. tropicalis isolates were assessed. The correlation between the virulence factors and antifungal susceptibility patterns was analyzed. During a 4year study, forty-five C. tropicalis isolates were recovered from candidemia patients. The isolates expressed different frequencies of virulence determinants as follows: coagulase 4 (8.9%), phospholipase 5 (11.1%), proteinase 31 (68.9%), esterase 43 (95.6%), hemolysin 44 (97.8%), biofilm formation 45 (100%) and CSH 45(100%). All the isolates were susceptible to amphotericin B and showed the highest resistance to voriconazole. There was a significant positive correlation between micafungin minimum inhibitory concentrations (MICs) and hemolysin production (rs = 0.316). However, we found a negative correlation between fluconazole MICs and esterase production (rs = -0.383). We observed the high expression of ERG11 and UPC2 genes in fluconazole-resistant C. tropicalis isolates. C. tropicalis isolated from candidemia patients extensively displayed capacities for biofilm formation, hemolysis, esterase activity, and hydrophobicity. In addition, the overexpression of ERG11 and UPC2 genes was considered one of the possible mechanisms of azole resistance.

Corynebacterium spp. - problematic pathogens of the human respiratory tract (review of literature).

Corynebacterium spp. - representatives of the normal microflora of the human body, but their role in the development of diseases in both immunocompromised and immunocompetent patients is known. Corynebacterim spp. (C. pseudodiphtheriticum, C. striatum, C. amycolatum, C. accolens, C. argentoratense, etc.) is associated with diseases of the respiratory tract: tracheitis, pharyngitis, rhinosinusitis, bronchitis, etc. They can be transmitted by airborne droplets, household contact, and possibly by hematogenic pathways. Corynebacterim spp. toxins do not produce, but are capable of adhesion and invasion, biofilm formation, production of neuraminidase, hyaluronidase, and hemolysin. It is necessary to take into account not so much the species, but the strain affiliation of isolates of Corynebacterium spp., since among the representatives of one species of non-diphtheria corynebacteria (for example, C. pseudodiphtheriticum), colonizing the respiratory tract, there may be strains that can exhibit not only pathogenic properties, but also probiotic activity. Microbiological diagnostics is based on their quantitative determination in biological material, phenotypic (culture study, test systems for biochemical identification, Vitek 2 automated systems) and genotypic (16SpRNA gene sequencing and rpoB) methods. It is possible to use mass spectrometric analysis (MALDI-ToF-MS). The greatest activity against Corynebacterium spp. in vitro studies preserve vancomycin, teicoplanin, and linezolid. Successful therapy with at least two of the following antimicrobial agents (AMP) has been reported: vancomycin, rifampicin, linezolid, and daptomycin. The sensitivity of isolates of Corynebacterium spp. to AMP is not related to the species, but is due to strain differences, and therefore it is necessary to test each isolated strain. Continuous monitoring of the sensitivity of Corynebacterium spp. strains to AMP is necessary due to the observed variability of these traits. Of particular importance is the identification of multidrug-resistant isolates that are currently considered highly pathogenic. When compiling the review, the databases Scopus, Web of Science, The Cochrane Library, CyberLeninka, RSCI were used.

Effect of Food Matrix Type on Growth Characteristics of and Hemolysin Production by Vibrio alginolyticus

The growth of and hemolysin production by two V. alginolyticus strains (HY9901 and ATCC 17749T) at 30°C were investigated in briny tilapia, shrimp, scallop, oyster, pork, chicken, freshwater fish, and egg fried rice. Bacteria were enumerated by plate counting. Hemolysin production was evaluated with blood agar and hemolytic titer tests. The two V. alginolyticus strains had similar growth and hemolysin production patterns in all tested foods. Based on the goodness-of-fit primary model statistics (coefficient of determination, mean square error, bias factor, and accuracy factor), the modified Gompertz model was a better fit than the logistic model to V. alginolyticus growth in foods. Growth kinetic parameters of V. alginolyticus had a higher μmax and shorter λ in the following order: briny tilapia > shrimp > freshwater fish > egg fried rice > scallop > oyster > chicken > pork. V. alginolyticus levels were similar at the stationary phase, with no significant growth difference between raw and cooked foods. Significantly higher thermostable direct hemolysin activity (P < 0.05) was found for V. alginolyticus in the following order: briny tilapia > freshwater fish > shrimp > chicken > egg fried rice > scallop > oyster > pork. However, the hemolytic titer was not consistent with the thermostable direct hemolysin activity and was significantly higher (P < 0.05) in the following order: briny tilapia > egg fried rice > shrimp > freshwater fish > chicken > scallop > oyster > pork. Contrary to current belief, V. alginolyticus produced more hemolysin in some nonseafoods (freshwater fish, egg fried rice, and chicken) than in scallops or oysters. This report is the first on the growth and toxicity of V. alginolyticus in different food matrices and confirms that some nonseafoods can be contaminated with pathogenic V. alginolyticus. These results should increase awareness of nonseafood safety issues and improve the accuracy of V. alginolyticus risk assessments.

Staquorsin: A Novel Staphylococcus aureus Agr-Mediated Quorum Sensing Inhibitor Impairing Virulence in vivo Without Notable Resistance Development.

The emergence of microbial resistance to the available antibiotics is a major public health concern, especially with the limited rate of developing new antibiotics. The utilization of anti-virulence agents is a non-conventional approach that can be used to combat microbial infection. In Staphylococcus aureus, many virulence factors are regulated by the Agr-mediated quorum sensing (QS). We developed a chemical compound that acts a potential Agr-inhibitor without reducing bacterial viability. The compound was designated staquorsin for Staphylococcus aureus QS inhibitor. In silico analyses confirmed the binding of staquorsin to the AgrA active site with an absolute binding score comparable to savirin, a previously described AgrA inhibitor. However, staquorsin turned out to be superior over savarin in not affecting the S. aureus viability in concentrations up to 600 μM. On the other hand, savirin inhibited S. aureus growth in concentrations as low as 25 μM. Moreover, staquorsin proved to be a potent inhibitor of the Agr system by inhibiting hemolysins, lipase production, and affecting biofilms formation and detachment. On the molecular level it significantly inhibited the effector transcript RNA III. In vivo testing, using the murine skin abscess model, confirmed the ability of staquorsin to modulate S. aureus virulence by effectively controlling the infection. Twenty passages of S. aureus in the presence of 40 μM staquorsin have not resulted in loss of activity as evidenced by maintaining its ability to reduce hemolysin production and RNA III transcript levels. In conclusion, we hereby describe a novel anti-virulence compound inhibiting the S. aureus Agr-system and its associated virulence factors. It is active both in vitro and in vivo, and its frequent use does not lead to the development of resistance. These findings model staquorsin as a promising drug candidate to join the fierce battle against the formidable pathogen S. aureus.

Candiduria—Study of Virulence Factors and Its Antifungal Susceptibility Pattern in Tertiary Care Hospital

Background The increased incidence of candiduria in hospitalized patients is due to the use of indwelling devices, long-term antibiotics, parenteral nutrition, and immunocompromised status of the patient. In this study, an attempt was made to speciate, characterize, and determine the antifungal susceptibility pattern of Candida isolated from urinary tract infections (UTIs). Materials and Methods A total of 70 Candida isolates were obtained from urine samples. The isolated Candida species were studied for the production of virulence factors like phospholipase, protease activities, hemolysin, and biofilm production. Antifungal susceptibility testing of the isolated yeasts was done using Mueller-Hinton agar supplemented with 0.5 mg/mL methylene blue by E-test method for amphotericin B, fluconazole, caspofungin, and voriconazole. Results Out of 70 isolates, Candida tropicalis was the most frequently isolated species (65.7%), followed by Candida albicans (14.3%), Candida glabrata (7.1%), Candida krusei (5.7%), Candida parapsilosis (4.3%), and Candida dubliniensis (2.9%). A total of 37.1% were biofilm producers, 62.9% showed proteinase activity, 38.6% were phospholipase positive, and 58.6% isolates showed hemolytic activity. Antifungal susceptibility profile of Candida species showed 38.6, 25.7, 15.7, and 12.9% resistance to amphotericin B, fluconazole, caspofungin, and voriconazole, respectively. Conclusion A rising trend in isolation of non-albicans Candida from urinary isolates was noticed, which was statistically significant when comparing catheterized and noncatheterized urinary isolates from our study. However, there was no statistically significant difference when different virulence factor expressions were compared among Candida spp. isolated from catheterized and noncatheterized urinary samples. Due to this rise in non-albicans Candida species causing UTI that are intrinsically resistant to certain antifungal agents like azoles and increasing incidence of antifungal resistance, it is essential to monitor the antifungal susceptibility profile of Candida species causing candiduria.

Urinary Tract Infections in Benin: Exploring the Virulence Factors and Antibiotic Resistance and Virulence Genes among Bacterial Isolates

Objective: The emergence and increasing spread of resistance to antibiotics in uropathogenic Enterobacteriaceae is a huge public health problem and increase the morbidity and mortality. The objective of this study was to produce scientific data on the virulence and level of resistance of these bacteria in Benin.&#x0D; Results: This study was conducted on 230 strains of enterobacteria isolated from patients with urinary tract infections. These virulence factors sought were hemolysis, hemagglutination, serum resistance, biofilm formation, and the production of lipase, protease and lecithinase. The molecular characterization of the virulence and antibiotic resistance genes was accomplished by PCR according to the different conditions for each gene. The bacteria possessed several types of virulence factors such as hemagglutinin (28.26%), lipase production (23.92%) and hemolysin production (17.83%). The virulence gene identified were FimH (74.79%), PapC (30.44%), Iss (11.34%) and Biofilm (3.92%). The antibiotic resistance genes such as Bla-SHV (49.56%), Bla-CTX-M15, Bla-CTX-M2 (33.92%), Bla-CTX-M9 (19.13%) and bla-CTX-M1 (11.74%) were also detected.&#x0D; Conclusion: These results filled the national gap about virulence and antimicrobial resistance of enterobacteria responsible of urinary tract infection and may be used to improve the management of patients in Benin.

Non-Candida albicans Candida species: virulence factors and species identification in India.

Background and Purpose: The predominant cause of candidiasis was Candida albicans which has recently changed to non-Candida albicans Candida (NCAC) (i.e., Candida spp. other than the C. albicans). The NCAC spp., earlier considered non-pathogenic or minimally virulent, are now considered a primary cause of morbidity and mortality in immunocompromised individuals. Given the NCAC spp.has become more common in clinical cases, this study aimed to determine the prevalence of NCAC spp. in different clinical specimens and assess a few of their virulence factors.Materials and Methods: Routine samples for bacterial culture and sensitivity that showed colony characteristics, like Candida on Blood Agar and microscopic features resembling Candida spp., were processed further. Candida isolates underwent tests for chlamydospore formation and biochemical tests, including sugar fermentation and sugar assimilation tests. These were grown at 42oC, and their colony color was identified using HiCromeTM Candida Differential Agar (HiMedia Laboratories Pvt. Ltd., Mumbai, India), HiCandidaTM Identification Kit (HiMedia Laboratories Pvt. Ltd., Mumbai, India), and VITEK-2® Compact (Biomérieux, France). Virulence factors, such as adherence to buccal epithelial cells (ABEC), biofilm formation, hemolytic activity, and production of coagulase enzyme were also tested.Results: Mean age of the patients was 38.46 years with a male-female ratio of 1.36:1. In total, 137 Candida isolates were recovered; 45.3%, 19.7%, and 13.9% of the isolates were isolated from urine, vaginal swabs, and oropharyngeal swabs, respectively. Moreover, 55 (40.1%) isolates were those of C. albicans and 82 (59.9%) isolates belonged to NCAC spp., with C. tropicalis (23.4%) contributing highest among NCAC species. Furthermore, C. albicans (3; 50%) was the most common spp. in cases of candidemia. Haemolysin production (85.5%) and ABEC (78.2%) were the major virulence factors in C. albicans. C. tropicalis (59.4%) and C. dubliniensis (50%) showed maximum ABEC. Biofilm forming capacity was higher in C. tropicalis (78.1%) than C. albicans (67%).Conclusion: Results of this study suggest varied prevalence and virulence based on geographical locations, even within a subcontinent. It clearly indicates the emergence of the NCAC spp. and their predominance in different body fluids. Identification of Candida to the spp. level should become a routine in all laboratories.

A novel anti-infective molecule nesfactin identified from sponge associated bacteria Nesterenkonia sp. MSA31 against multidrug resistant Pseudomonas aeruginosa

Overuse of antibiotics coupled with biofilm-forming ability has led to the emergence of multi-drug P. aeruginosa strains worldwide. Quorum sensing is a bacterial cell-cell communication system that regulates the expression of genes, including virulence factors, through production of acyl-homoserine lactones (AHLs) in Pseudomonas aeruginosa. The phenotypic expression of virulence factors in P. aeruginosa is mediated by quorum sensing systems (las and rhl). In this study an anti-infective molecule produced by a marine actinomycetes Nesterenkonia sp. MSA31 was elucidated as lipopeptide by NMR and LC-MS/MS analysis. The new lipopeptide molecule was named Nesfactin. This molecule effectively inhibited virulence phenotypes including production of hemolysin, protease, lipase, phospholipase, esterase, elastase, rhamnolipid, alginate, and pyocyanin, as well as motility and biofilm formation in P. aeruginosa. The high-performance thin layer chromatography (HPTLC) analysis revealed that the lipopeptide (50 μg/mL) inhibited production of the AHLs produced by the las and rhl quorum sensing systems (3-oxo-C12-HSL and C4-HSL, respectively). Docking analysis showed the binding affinity of the ligand towards the quorum sensing receptor molecules. The confocal laser scanning microscopy images showed the anti-biofilm effect of lipopeptide against P. aeruginosa. Nesfactin based hydrogel showed a significant antibiofilm effect on the catheter. This study suggests that the lipopeptide may be an effective anti-virulence treatment for Pseudomonas aeruginosa infections.

Possible antivirulent activity of some agents against clinical isolates of Pseudomonas aeruginosa

Background: Pseudomonas aeruginosa has developed different mechanisms of resistance against antibiotics and became one of the most life-threatening pathogens. Fighting against its virulence Factors are an alternative therapeutic target. Objective: This study was directed towards the investigation of anti-quorum sensing activity and inhibitory action on virulence factors of different agents including antibacterial agents to which Pseudomonas aeruginosa isolates are resistant and non-antibacterial agents. Methodology: Anti-quorum sensing activity of ceftriaxone, ceftazidime (CAZ), cefepime (FEP), vancomycin (VA), paracetamol (PA), and pheniramine maleate (PHE) investigated as well as their ability to reduce other virulence factors including protease, hemolysin, and pyocyanin production. Results: This study showed that 3rd and 4th generations cephalosporins could be used as anti-quorum sensing agents effectively in the treatment of Pseudomonas aeruginosa infections, however, vancomycin, paracetamol, and pheniramine maleate had no effect on inhibiting the studied virulence factors. Conclusion: From our study we conclude that although cephalosporins at the used concentrations did not show anti-pseudomonal activity they were effective as anti virulent agents that could be utilized in therapeutically in controlling Pseudomonas aeruginosa infections.

Analysis of phenotypic and genotypic methods for determining the biofilm-forming abilities of CoNS isolates: Association with hemolysin production and the bacterial insertion sequence elements IS256/257

Biofilm-forming Coagulase-negative staphylococci (CoNS) has been considered the etiological agent in various infections. The purpose of this study was to determine the biofilm-producing ability and to explore the virulence factors associated with biofilm formation in these isolates. Ninety-one isolates of CoNS were characterized and subjected to biofilm detection by the microtiter plate (MTP) and Congo red agar (CRA). The presence of biofilm-associated genes, insertion sequence elements IS256 and IS 257, and genes responsible for producing hemolytic toxins in CoNS isolates were screened using the PCR method. Out of the 91 clinical isolates of CoNS, 49 (53.8), 37 (40.7%), and 5 (5.5%) isolates were identified as S. epidermidis, S. haemolyticus, and S. saprophyticus, respectively. The prevalence of biofilm-associated genes icaA, icaB, icaC, icaD, atlE and bhp in CoNS was: 27.5%, 16.5%, 21%, 24.2%, 52% and 1% respectively. The incidence of hemolysin-encoding genes was as follows; hla (88%), hla_yiD (63%), hlb (47.3%), and hld (56%). Furthermore, the occurrence of IS256/IS257 and the capability to form a biofilm. In conclusion, high rates of hemolysin-producing CoNS isolates were detected in both biofilm-forming and resistant strains. Thus, the co-existence of hemolysin and biofilm-associated genes play an important role in emerging violent CoNS bacteria. The prevalence of hemolysin-producing CoNS isolates also suggests the presence of other virulence factors. However, we believe that the explanation for ica operon being more associated with the hemolysin production in this study could be the current investigation's geographical location. Due to the hemolysin production and biofilm formation correlation, it is necessary to study epidemiology and identify these factors.

Back to Nature: Combating Candida albicans Biofilm, Phospholipase and Hemolysin Using Plant Essential Oils.

Candida albicans is the causative agent of fatal systemic candidiasis. Due to limitations of antifungals, new drugs are needed. The anti-virulence effect of plant essential oils (EOs) was evaluated against clinical C. albicans isolates including cinnamon, clove, jasmine and rosemary oils. Biofilm, phospholipase and hemolysin were assessed phenotypically. EOs were evaluated for their anti-virulence activity using phenotypic methods as well as scanning electron microscopy (SEM) and atomic force microscopy (AFM). Among the C. albicans isolates, biofilm, phospholipase and hemolysins were detected in 40.4, 86.5 and 78.8% of isolates, respectively. Jasmine oil showed the highest anti-biofilm activity followed by cinnamon, clove and rosemary oils. SEM and AFM analysis showed reduced adherence and roughness in the presence of EOs. For phospholipase, rosemary oil was the most inhibitory, followed by jasmine, cinnamon and clove oils, and for hemolysins, cinnamon had the highest inhibition followed by jasmine, rosemary and clove oils. A molecular docking study revealed major EO constituents as promising inhibitors of the Als3 adhesive protein, with the highest binding for eugenol, followed by 1,8-cineole, 2-phenylthiolane and cinnamaldehyde. In conclusion, EOs have a promising inhibitory impact on Candida biofilm, phospholipase and hemolysin production, hence EOs could be used as potential antifungals that impact virulence factors.

Virulence Genes and Resistance Profile of Escherichia coli Isolated in Meat Meal Samples

Background: Feed is the main route of transmission of pathogenic microorganisms and is responsible for a large part of the cost of poultry production, so the inclusion of alternative foods in diets for monogastrics has been a constant. Among alternative foods most used in modern poultry farming are animal meal, however, when contaminated they constitute a route of transmission of several pathogenic agents, including Escherichia coli. In addition, there is a zoonotic potential, as poultry products are intended for human consumption. The objective of this research was to detect virulence genes, as well as to evaluate the resistance profile of Escherichia coli isolates from meat meal samples. Materials, Methods &amp; Results: A total of 40 Escherichia coli isolates were analyzed and the virulence genes surveyed iss, ompT, hlyF, iutA, and fimA identified by Polymerase Chain Reaction (PCR). The antimicrobial agents tested were: amoxycillin (30 μg), ceftiofur (30 μg), ciprofloxacin (5 μg), doxycycline (30 μg), florfenicol (30 μg), fosfomycin (200 μg), gentamicin (10 μg), norfloxacin (10 μg) and oxacillin (1 μg). It was possible to observe the occurrence of the iss resistance gene in 100% of Escherichia coli isolates, followed by hlyF (85%), fimA (75%), ompT (17.5%) and iutA (5%). Regarding the simultaneous detection for the genes, a greater association between the genes iss, hlyF and fimA (60%) was verified. All isolates showed resistance to oxacillin (100%), followed by doxycycline (25%), amoxicillin (22.5%), norfloxacin (17.5%), ceftiofur (15%), florfenicol (12.5%), fosfomycin (12.5%), ciprofloxacin (10%) and gentamycin (2.5%). In this study, a variation of the multiple antimicrobial resistance index (IRMA) was observed between 0.22 and 0.77. Discussion: The indiscriminate use of of antimicrobials as performance enhancers in production animals, may have contributed to the increase in antimicrobial resistance, with the occurrence of multiresistant Escherichia coli carrying virulence genes. Virulence genes present in Escherichia coli isolates are studied to understand the degree of influence they exert in the establishment of the disease, one of the most researched genes is the iss gene, involved in the processes that promote serum resistance. In this study, iss (100%) was present in all the isolates analyzed, although it is not the only mechanism used by these bacteria to reach internal organs and trigger an infection, this gene encodes an important mechanism associated with high levels of virulence. The second highest prevalence found was of the hlyF gene (85%), the high prevalence of hlyF suggests virulence potential, involved with the production of hemolysin and improvement of outer membrane vesicles associated with the release of toxins. The fimA gene (75%) was detected in a slightly lower percentage when compared to iss and hlyF. With the second lowest prevalence, the ompT gene (17.5%), is involved in a process that includes the proteolytic degradation of antimicrobial peptides and with the lowest prevalence the iutA gene (5%). Certain combinations of virulence genes make the strains easier to survive, adhere to, colonize and even the ability to develop septicemic conditions. Multiresistant E. coli strains, is a fact of concern for both animal and human health, since the presence of multiresistant strains, originating from poultry, can be transmitted from chicken carcasses. In this sense, the importance of sanitary control of the inputs used in animal feed is emphasized, as well as the prudent use of antimicrobials in animal production, with a view to producing a safe food, minimizing not only the economic losses, but also the risks to human health. Keywords: antimicrobial, bacterial resistance, colibacillosis, poultry.

Pattern in Minimum Inhibitory Concentration of Vancomycin and Tigecycline on Clinical Isolates of Enterococcus Species Isolated from Clinical Samples at our Tertiary Care Hospital

Background: Genus Enterococcus have emerged as nosocomial pathogens. Urinary tract infections followed by intra-abdominal abscesses, Wounds, usually intra-abdominal or pelvic, are the next most common sites. Bacteremia is the third most common infection caused by Enterococci. Enterococcal meningitis is rare and is seen primarily in neonates and inpatients who have undergone complicated neurosurgical procedures [4]. There is an increasing role of Enterococci in nosocomial infections in the recent years mostly because of their intrinsic resistance to many clinically useful antibacterial drugs. One of the leading cause of nosocomial infection is Vancomycin-resistant Enterococci (VRE). For good therapy an effective, accurate and early detection of VRE along with Minimum Inhibitory Concentrations (MIC) is necessary. Tigecycline has a broad spectrum of activity against enterococci (vancomycin-susceptible). Haemolysin is a cytolytic protein and the producing strains of Enterococci have been shown to be associated within creased severity of infection [9]. In order to know the current scenario of minimum inhibitory concentration of Vancomycin and Tigecycline and the production of Haemolysin by some strains of Enterococci, the present study has been done. Methods: This study was performed on Clinical isolates of Enterococcus which were received from tertiary care hospitals and processed at Department of Microbiology, Kasturba Medical College, Mangalore and Microbiology diagnostic laboratory (KMC hospital, Ambedkar circle), Mangalore. The study was done for a period of 6 months from Nov 2017-May2018. Convenient non-random sampling method was followed to collect clinical isolates. All isolates of Enterococcus from various clinical specimens like pus, deep tissue, urine, blood and body fluids were included in the study. Result: Out of 100 Enterococcus isolates collected, majority were of E. faecium. Among all the isolates, 5% were resistant to vancomycin, but 100% sensitivity to tigecycline. Hemolysin produced by 46% of the total isolates. It has shown its association of the Enterococcus species to various antibiotics and their resistant pattern.

Study of Virulence Factors and Antimicrobial Susceptibility Pattern of Uropathogenic Escherichia Coli in a Tertiary Care Hospital

Introduction: Urinary tract infections are the most commonly encountered bacterial infections both in community and health care settings. Uropathogenic Escherichia coli (UPEC) is the single most common pathogen, accounting for 70-75% of all cases of Urinary Tract Infections (UTIs). The aim of the present study is to isolate UPEC from urine samples from UTI patients, to evaluate the virulence factors and antimicrobial susceptibility pattern. Material and methods: A total of 100 E.coli isolates out of 292 urine samples collected from clinically suspected UTI were taken for the study and detection of virulence factors such as haemolysin production, haemagglutination and cell surface hydrophobicity were done. Antibiotic susceptibility test was done by Kirby Baur disc diffusion method as per the CLSI guidelines. Results: The incidence of UTI was more in females 52% and haemolysis was seen in 32%, haemagglutination in 40%, and cell surface hydrophobicity in 24% of strains. The UPEC strains were more resistant to Ampicillin (52%) followed by Cotrimoxazole (46%), Norfloxacin (42%) and Ceftriaxone(40%). Most of the strains were sensitive to Meropenem (80%) followed by Amikacin(78%) and Nitrofurantoin (72%). Conclusion: As there is a significant association between the virulence and antimicrobial resistance of UPEC, a routine testing of these factors is recommended and further studies at molecular level are necessary.

&amp;lt;i&amp;gt;In Vitro&amp;lt;/i&amp;gt; Efficacy of Clove Oil and Eugenol against &amp;lt;i&amp;gt;Staphylococcus&amp;lt;/i&amp;gt; spp and &amp;lt;i&amp;gt;Streptococcus mutans&amp;lt;/i&amp;gt; on Hydrophobicity, Hemolysin Production and Biofilms and their Synergy with Antibiotics

The present study aimed to evaluate Syzygium aromaticum (clove) plant extract, clove oil and eugenol for their antibacterial activity and their potential to eradicate bacterial biofilms alone and in combination with antibiotics. Anti-bacterial efficacy of S. aromaticum extract, clove oil and eugenol was evaluated as minimum inhibitory concentration (MIC) and subsequently sub-MICs was selected for inhibition of virulence factors against test bacterial strains. Biofilm cultivation and eradication was assayed using XTT reduction in 96-well microtiter plate. Checkerboard method was used to study the interaction between essential oils and antibiotics. Staphylococcus aureus MTCC3160, Staphylococcus epidermidis MTCC435, Staphylococcus sciuri (SC-01), Staphylococcus auricularis (SU-01) and Streptococcus mutans MTCC497 were found strong biofilm former among all the test bacterial strains. The potency of test agents was found in the order of eugenol > clove oil > S. aromaticum methanolic extract. Sub-MIC (0.5 × MIC) of clove oil and eugenol showed a significant reduction in cell surface hydrophobicity (p S. auricularis (SU-01), S. epidermidis MTCC435 and S. mutans MTCC497 compared to planktonic MIC (PMIC). Antibiotics (vancomycin and azithromycin) exhibited upto 1000-folds increased in SMIC compared to PMIC against all the test bacterial strains. Synergy was observed between eugenol and antibiotics (vancomycin/azithromycin) against all the test bacterial strains in both planktonic and sessile mode. Highest synergy was exhibited between eugenol and azithromycin in planktonic mode (FICI value 0.141). Further, microscopy also confirmed the spectacular effect of combination treatment on pre-formed S. aureus MTCC3160 and S. mutans MTCC497 biofilms. These findings highlighted the promising role of clove oil and eugenol alone and in combination on pathogenic bacterial biofilms.