Interaction of Salmonella with E. coli and Proteus spp. in Biofilm Formation

Aims: Investigate the interaction of Salmonella spp. with E. coli and Proteus spp. in biofilm formation as mono and dual-species at different time durations Experimental Design: Salmonella, Proteus, and E. coli were isolated from Broiler chicken meat, and the biofilm-forming ability of these organisms were studied. Place and Duration of Study: The study was conducted at the Laboratory of Livestock Production, Faculty of Agricultural Sciences, Sabaragamuwa University of Sri Lanka, from 2019 December to 2020 May. Methodology: This study investigated the biofilm-forming ability of Salmonella as a mono species and its interaction with E. coli and Proteus in the process of biofilm formation. Microorganisms used for this study were isolated from broiler chicken meat. Biofilm was quantified using a microtitre plate assay. The interaction effects were tested at the temperature of 280C in different time durations (up to 120 hours). Results: Salmonella 1 and Proteus monocultures showed significantly higher biofilm-forming ability than Salmonella 3 isolate at all tested time points. At 120 hr, additionally to the salmonella 1 and Proteus isolates E. coli also formed significantly higher biofilms than Salmonella 3. However, Salmonella 3 was the lowest biofilm former as mono biofilm at all tested time durations. Salmonella 1 interaction with Salmonella 3 isolates formed less biofilms than Salmonella 1 mono biofilm at 48hr and 72hr correspondingly. Salmonella 1 and its interactions with Salmonella 3, Proteus, E. coli showed similar biofilm-forming abilities without significant differences at all other tested time points. Specifically, Salmonella 3 interaction with Salmonella 1 as dual biofilm showed higher biofilm-forming ability than Salmonella 3 mono biofilm at all tested time points. Tested isolates and their interaction achieved the highest biofilm formation at numerous time points. In fact, at 48hr, Salmonella 3 isolates and its interaction of Proteus, E. coli, and Salmonella 1 interaction with Proteus attained their highest biofilm formation abilities. The highest biofilm formation was achieved by Salmonella 1 isolate as mono biofilm and Salmonella 1 interaction with E. coli as dual biofilm at 72hr. Biofilm-forming trend of respective isolates and interactions showed numerous patterns at tested time durations. Specifically, E. coli rapidly enhanced its biofilm-forming ability as monoculture from 24 hr to 120 hr. Proteus, Salmonella 3 as monocultures, Salmonella 3 interaction with Proteus and E. coli as dual cultures showed progressive biofilm development from 24 hr to 48 hr. Salmonella 1 monoculture and its interaction with Salmonella 3, E. coli as dual biofilm improved their biofilm-forming ability from 24 hr to 72 hr. Similar to Salmonella 3 interaction with Proteus, Salmonella 1 interaction with Proteus also increased its biofilm-forming ability from 24 hr to 48 hr. Conclusions: This study concluded that there is a variation among isolates and their combinations in forming the biofilms, where there is an enhancement of biofilm in dual-species over the mono-species in some interaction, and there is a reduction in biofilm formation by dual-species with some combinations. Further, this concluded that Salmonella is interacting with other commonly found bacteria such as Proteus and E. coli in biofilm formation.

Detection of catheter related blood stream infections in an ICU of a tertiary care center in Northern India

Indwelling medical devices are frequently used in all health setting with critical care units of hospitals for treatment and intervention in patient care.Microorganisms attach to surfaces resulting in the formation of a biofilm which pose a serious public health issue because of its increased resistance to antimicrobial agents and the potential to cause infections.To determine the proportion of bacterial Biofilms in patients with central venous line and to find out the organisms most commonly associated with it.The study was carried out from January 2019 to September 2020. Positive cultures were obtained from 58 of these 102 samples. The isolates were then tested for in vitro production of biofilm using a microtitre plate assay. All the isolates were subjected to antimicrobial susceptibility tests on Muller-Hinton agar by Kirby Bauer disc diffusion method based on CLSI guidelines. Demographic characteristics of the study subjects showed that among 102 catheterized patients isolated with males 58.82% and 41.1% females. Out of 58 isolates, 62.06% were gram negative bacteria (GNB) and 37.93% were gram positive. was most common isolate 27.58% followed by 20.68%, 15.5%, respectively. The total number of positive slime producers, in this study, was 23(39.6%). The highest number of strong slime producer strains was observed in case of (2 out of 5) (4 out of 12) and 3 out of 16)Antibiotic susceptibility patterns showed increased resistance towards penicillin and beta -lactum group of antibiotics, increased sensitivity to linezolid and vancomycin among gram positive organisms. Among gram negative bacteria increased resistance was seen for cephalosporins and aminigylycosides and least resistance for colistin.Colonization of indwelling medical devices with consequent biofilm production is a likely contributory factor to infections. The microorganisms survive in the hospital environment despite unfavourable conditions such as desiccation, nutrient starvation, and antimicrobial treatment. Resistance to antibiotics ladder is increasing and it’s necessary to take actions to reduce its hindrance in the future. Advanced studies in biofilm will help to prevent the more virulent factors which protect the bacteria from host immunity and to get rid of critical complications in therapy.

Evaluation of the role of staphylococci in the pathomechanism of conjunctivitis

Purpose Determination of the association between ica genes and phenotypic biofilm formation in staphylococcal isolates involved in conjunctivitis, their antibiotic resistance as well as detection of selected virulence characteristics: adhesion to epithelial cells and in vitro cytotoxicity. Methods The study included 26 Staphylococcus aureus (SA) and 26 Staphylococcus epidermidis (SE) isolates. The presence of icaAD genes and ica operon was determined by the PCR assay. Phenotypic biofilm formation was verified using the microtitre plate assay. Antibiotic resistance was performed using the disc diffusion method. Staphylococcal ability to attach to host cells was assessed by flow cytometry. Cytotoxicity on epithelial cells was evaluated by LDH assay. Results The ica genes were detected in 26.9% of SE and in 42.3% of SA isolates. Only 15.3% of isolates (SE) were positive for both the icaAD and the ica operon. Phenotypically, 19.2% of SE isolates were strong biofilm producers, among which three were both icaAD- and ica operon-positive. About 26.9% of SA isolates were strong biofilm producers. Methicillin resistance (MR) was detected in 34.6% of SE and 26.9% of SA isolates. About 75% of MR isolates were multidrug resistant. SA isolates adhered to host cells more extensively than SE. SA isolates released higher level of LDH than SE. Conclusions Adherence abilities were commonly observed in staphylococci associated with conjunctivitis. However, low prevalence of isolates positive for a complete and functional ica locus and low prevalence of strong biofilm producers was detected. SA adhered to a greater extent to eukaryotic cells than SE and were more cytotoxic.

Carprofen elicits pleiotropic mechanisms of bactericidal action with the potential to reverse antimicrobial drug resistance in tuberculosis

Background The rise of antimicrobial drug resistance in Mycobacterium tuberculosis coupled with the shortage of new antibiotics has elevated TB to a major global health priority. Repurposing drugs developed or used for other conditions has gained special attention in the current scenario of accelerated drug development for several global infectious diseases. In a similar effort, previous studies revealed that carprofen, a non-steroidal anti-inflammatory drug, selectively inhibited the growth of replicating, non-replicating and MDR clinical isolates of M. tuberculosis. Objectives We aimed to reveal the whole-cell phenotypic and transcriptomic effects of carprofen in mycobacteria. Methods Integrative molecular and microbiological approaches such as resazurin microtitre plate assay, high-throughput spot-culture growth inhibition assay, whole-cell efflux inhibition, biofilm inhibition and microarray analyses were performed. Analogues of carprofen were also synthesized and assessed for their antimycobacterial activity. Results Carprofen was found to be a bactericidal drug that inhibited mycobacterial drug efflux mechanisms. It also restricted mycobacterial biofilm growth. Transcriptome profiling revealed that carprofen likely acts by targeting respiration through the disruption of membrane potential. The pleiotropic nature of carprofen’s anti-TB action may explain why spontaneous drug-resistant mutants could not be isolated in practice. Conclusions This immunomodulatory drug and its chemical analogues have the potential to reverse TB antimicrobial drug resistance, offering a swift path to clinical trials of novel TB drug combinations.

Distribution of genes encoding adhesins and biofilm formation capacity among Uropathogenic Escherichia coli isolates in relation to the antimicrobial resistance

Background: Escherichia coli is the most predominant pathogen involved in UTIs. Mainly, fimbrial surface appendages are impli- cated in adherence to urothelium besides non-fimbrial proteins. Objectives: To determine prevalence of genes encoding fimbrial and non-fimbrial proteins among Uropathogenic Escherichia coli (UPEC). Furthermore, distribution of these genes and biofilm formation capacity were investigated in relation to antimicrobial resistance. Methods: Antimicrobial susceptibility of 112 UPEC isolates was performed using disc diffusion method. ESBL production was confirmed by double disc synergy test. Genes encoding fimbrial and non-fimbrial proteins were detected using PCR and biofilm formation was investigated using microtitre plate assay. Results: UPEC isolates exhibited high resistance against doxycyclines (88.39 %), β-lactams (7.14-86.6%), sulphamethoxaz- ole–trimethoprim (53.75%) and fluoro-quinolones (50%). Fifty percent of tested isolates were ESBL producers. PapGII gene was statistically more prevalent among pyelonephritis isolates. SfaS, focG and picU genes were statistically associated with flu- oro-quinolone (FQs) sensitive isolates and Dr/afaBC gene was statistically associated with ESBL production. Moreover, non- MDR isolates produced sturdier biofilm. Conclusion: PapGII adhesin variant seems to have a critical role in colonization of upper urinary tract. There is a possible link between antimicrobial resistance and virulence being capable of affecting the distribution of some genes besides its negative impact on biofilm formation. Keywords: Urinary tract infection; Escherichia coli; UPEC; adhesin genes; ESBL; biofilm.

Detection of enterotoxin gene (sea) and biofilm formation ability among multidrug resistant Staphylococcus aureus isolated from shawarma sandwich sold at selected kiosks in Klang Valley, Malaysia

The occurrence of multi-drug resistant Staphylococcus aureus in food product of animal origin has increased the concern about their spread into the food supply chain. Presence of multidrug-resistant S. aureus in food products, including ready-to-eat foods imposes potential hazard for consumers. The objective of this research was to investigate the presence of multi-drug resistance of S. aureus in sixty ready-to-eat shawarma sandwiches. Agar-disc diffusion assay determined their resistance to 11 antibiotics. The sea and sed enterotoxin genes were detected by polymerase chain reaction method. Biofilm formation potential (BFP) was quantified by microtitre plate assay. The result revealed that thirty-six samples (60%) were positive for S. aureus. Majority of the isolates (n = 29; 80.6%) were resistant to at least one antibiotic. The isolates demonstrated highest resistance against ampicillin (69.4%) and penicillin (69.4%), while resistance to ciprofloxacin, tetracycline and kanamycin were 47.2%, 33.3% and 22.2%, respectively. Several isolates were resistant to trimethoprim (5.6%), trimethoprim-sulfamethoxazole- (2.8%), gentamicin (2.8%) and cephalothin (2.8%), while none exhibited resistance to chloramphenicol and nitrofurantoin. Out of the thirty-six isolates, twelve isolates (33.3%) were resistant to three or more classes of antibiotic (multidrug-resistant) and 50% had a Multiple Antibiotic Resistance index value more than 0.25. Of the multi-drug resistant isolates, four were positive for sea genes but no sed genes were present. All multi-drug resistance isolates were biofilm formers with five and six isolates were strong and moderate formers, respectively. Additionally, all the sea gene carrying multi-drug resistance isolates were strong biofilm formers. These findings revealed shawarma as a potential vehicle for the spread of multidrug-resistant S. aureus, suggesting more control measures for ready-to-eat food.

Antibiogram and Biofilm Formation among Carbapenem Resistant Klebsiella pneumoniae

Objectives: This cross-sectional study was designed to detect the carbapenemase producing K. pneumoniae along with biofilm producers from different clinical specimens and to compare antibiotic susceptibility pattern of biofilm producing carbapenem resistant Klebsiella pneumoniae and biofilm non-producing carbapenem resistant Klebsiella pneumoniae. Methods: A total of 1475 non-repetitive clinical samples were included on this study. Antibiotic Sensitivity Testing (AST), Modified Hodge Test (MHT) and Modified Carbapenem inactivation method (mCIM) were performed for detection of carbapenemase production and Congo red agar method (CRA) along with Microtitre plate method were performed for detecting biofilm production. Results: Among the clinical specimens cultured, growth positivity was 62.71%. E. coli was most predominant organism followed by K. pneumoniae (17.89%). Among the 110 K. pneumoniae, 57 were found to be carbapenemase producer. Majority of the carbapenemase producing K. pneumoniae were isolated from sputum (45.61%), in the specimen collected from age group 61-70 (28.07%) and in out-patient department (50.88%). Similarly, 65.45% K. pneumoniae out of 110 were found to be biofilm producer by Congo red agar method while among those 72, 73.59% isolates were found to be quantitatively biofilm producer in Microtitre plate assay. Out of 57 carbapenemase producer, 35.08% were strongly biofilm producer while among 53 carbapenemase non-producer 30.18% were strongly biofilm producer from Congo red agar method. Moreover, Microtitre plate assay evidenced that, out of 57 carbapenemase producer, 40.35% were highly biofilm producing and among the 15 carbapenemase nonproducer 66.66% were highly biofilm producer. Conclusion: Biofilm formation is highly prevalent with varying degree of resistance among different antibiotics including carbapenems that further augments antibiotic resistance. The study showed carbapenemase producers are stronger biofilm producer than the non-carbapenemase producer. Therefore, it is recommended to identify biofilm formation among carbapenemase producers for effective choice of antibiotics.

Antimicrofouling activities of marine macroalga Dictyota dichotoma from the Red Sea

Introduction. Marine organisms produce a variety of secondary metabolites mainly for achieving the defence against the competitors and predators. These compounds could be used as natural product antifoulants for the management of biofouling growth on marine structures. Objectives. To understand the antifouling defence strategies of marine macroalgae collected from the Red Sea. Methodology. The macroalga Dictyota dichotoma was collected from the Obhur Creek of Red Sea, Saudi Arabia and extracted using methanol. Surface and total extraction methods were performed and tested against a bacterial strain isolated from the microfouling assemblage. Results. The extracts obtained from the macroalgal samples have strong antibacterial and antibiofilm activities against the bacterial strain isolated from the marine microfouling assemblage. The total extracts showed strong bacterial growth inhibitory activities in culture plate method. In microtitre plate assay, surface extract showed strong biofilm inhibitory activity. GC-MS analysis indicated considerable variations in the metabolic profile of the surface and total extracts. Conclusion. This study revealed the importance of surface-associated compounds in antifouling defence mechanism of the marine macroalgae.

Antimicrofouling activities of marine macroalga Dictyota dichotoma from the Red Sea

Introduction. Marine organisms produce a variety of secondary metabolites mainly for achieving the defence against the competitors and predators. These compounds could be used as natural product antifoulants for the management of biofouling growth on marine structures. Objectives. To understand the antifouling defence strategies of marine macroalgae collected from the Red Sea. Methodology. The macroalga Dictyota dichotoma was collected from the Obhur Creek of Red Sea, Saudi Arabia and extracted using methanol. Surface and total extraction methods were performed and tested against a bacterial strain isolated from the microfouling assemblage. Results. The extracts obtained from the macroalgal samples have strong antibacterial and antibiofilm activities against the bacterial strain isolated from the marine microfouling assemblage. The total extracts showed strong bacterial growth inhibitory activities in culture plate method. In microtitre plate assay, surface extract showed strong biofilm inhibitory activity. GC-MS analysis indicated considerable variations in the metabolic profile of the surface and total extracts. Conclusion. This study revealed the importance of surface-associated compounds in antifouling defence mechanism of the marine macroalgae.