This study was undertaken to investigate the influence of culture conditions and medium components on production of antibacterial compounds by Serratia sp. WPRA3 (JX020764) which was isolated from marine water of Port Dickson, Malaysia. Biochemical, morphological, and molecular characteristics suggested that the isolate is a new candidate of the Serratia sp. The isolate showed strong antimicrobial activity against fungi, Gram-negative and Gram-positive bacteria. This bacterium exhibited optimum antibacterial compounds production at 28°C, pH 7 and 200 rev/min aeration during 72 h of incubation period. Highest antibacterial activity was obtained when sodium chloride (2%), yeast extract (0.5%), and glucose concentration (0.75%) were used as salt, nitrogen, and carbon sources respectively. Different active fractions were obtained by Thin-Layer Chromatography (TLC) and Flash Column Chromatography (FCC) from ethyl acetate crude extracts namely OCE and RCE in different culture conditions, OCE (pH 5, 200 rev/min) and RCE (pH 7/without aeration). In conclusion, the results suggested different culture conditions have a significant impact on the types of secondary metabolites produced by the bacterium.
Quorum-sensing regulates bacterial biofilm formation and virulence factors, thereby making it an interesting target for attenuating pathogens. In this study, we investigated anti-biofilm and anti-quorum-sensing compounds from secondary metabolites of halophiles marine streptomyces against urinary catheter biofilm forming Proteus mirabilis without effect on growth viability. A total of 40 actinomycetes were isolated from samples collected from different places in Iraq including marine sediments and soil samples. Fifteen isolates identified as streptomyces and their supernatant screened as anti-quorum-sensing by inhibiting quorum-sensing regulated prodigiosin biosynthesis of Serratia marcescens strain Smj-11 as a reporter strain. Isolate Sediment Lake Iraq (sdLi) showed potential anti-quorum-sensing activity. Out of 35 clinical isolates obtained from Urinary catheter used by patient at the Universiti Kebangsaan Malaysia Medical Center, 22 isolates were characterized and identified as Proteus mirabilis. Isolate Urinary Catheter B4 (UCB4) showed the highest biofilm formation with highest resistance to used antibiotic and was chosen for further studies. Ethyl acetate secondary metabolites extract was produced from sdLi isolate. First, we determined the Minimum Inhibitory Concentration (MIC) of sdLi crude extract against UCB4 isolate, and all further experiments used concentrations below the MIC. Tests of subinhibitory concentrations of sdLi crude extract showed good inhibition against UCB4 isolate biofilm formation on urinary catheter and cover glass using Scanning electron microscopy and light microscopy respectively. The influence of sub-MIC of sdLi crude extract was also found to attenuate the quorum sensing (QS)-dependent factors such as hemolysin activity, urease activity, pH value, and motility of UCB4 isolate. Evidence is presented that these nontoxic secondary metabolites may act as antagonists of bacterial quorum sensing by competing with quorum-sensing signals for receptor binding.
Microbial communities of two oil reservoirs from Malaysia, denoted as Platform Bo and Platform Pe were studied using
culture-independent approach. Environmental DNA was extracted and the universal amplified ribosomal region (UARR)
was target amplified for both prokaryotes and eukaryotes. The amplified products were purified and cloned into pTZ57R/T
vector to construct the 16S/18S rDNA library. Restriction endocucleases HhaI and MspI were used to screen the library.
From that, 125 and 253 recombinant plasmid representative clones from Platform Bo and Platform Pe, respectively, were
sent for DNA sequencing. Twenty-six operational taxonomic units (OTUs) consist of 20 genera detected at Platform Bo
and 17 OTUs consist of 13 genera detected at Platform Pe. Marinobacter and Acinetobacter species co-occurred in both
platforms whereas the rest are site-specific. Gammaproteobacteria accounted for 86.0% of the microbial community in
Platform Bo, where OTUs affiliated to Marinobacter, Pseudomonas and Marinobacterium that were the most abundant. The
major OTUs in the Platform Pe were with affinities to Achromobacter, followed by Stenotrophomonas and Serratia. The
only archaeal isolates were detected in Platform Pe, which affiliated to Thermocladium. The singletons and doubletons
accounted for about 50.0% of the OTU abundance in both platforms, which considerably significant despite their rare
occurrence.
In this work we report on the isolation of a local molybdenum-reducing bacterium. The bacterium reduced molybdate or Mo(6+) to molybdenum blue (oxidation states between 5+ to 6+). Electron donors that supported cellular growth were sucrose, maltose, mannitol, fructose, glucose and starch (in decreasing order) with sucrose supporting formation of the highest amount of molybdenum blue at 10 g/l after 24 hours of static incubation. The optimum molybdate and phosphate concentrations that supported molybdate reduction were 20 and 5 mM, respectively. Molybdate reduction was optimal at 37 degrees C. The molybdenum blue produced from cellular reduction exhibited a unique absorption spectrum with a maximum peak at 865 nm and a shoulder at 700 nm. The isolate was tentatively identified as S. marcescens strain Dr.Y9 based on carbon utilization profiles using Biolog GN plates and partial 16S rDNA molecular phylogeny. No inhibition of molybdenum-reducing activity was seen using electron transport system (ETS) inhibitors such as antimycin A, 1HQNO (Hydroxyquinoline-N-Oxide), sodium azide and cyanide suggesting that the ETS of this bacterium is not the site of molybdate reduction.
The success of microbial termiticides in controlling termites depends on the ability of microbes to grow in different media and the functionality of the microbes as a resistant barrier or toxic bait. This study was conducted to understand the mortality rate and behaviour changes of the subterranean termite Coptotermes curvignathus Holmgren introduced with different concentrations of Serratia marcescens strain LGMS 1 and Pseudomonas aeruginosa strain LGMS 3 using wood and soil as bacterial transfer medium. In general, higher concentration of bacteria in soil caused a reduction in tunnelling activity and wood consumption and an increase in mortality. However, application on wood revealed a different outcome. Wood treated with S. marcescens of 106 CFU ml-1 concentration proved to be more efficient as bait than higher concentration applications as it caused a high mortality rate while still highly palatable for termites. Wood or soil treated with S. marcescens concentration higher than 109 CFU ml-1 creates a high toxicity and repellent barrier for termites. Pseudomonas aeruginosa of 109 CFU ml-1 concentrations applied on wood served as a slow-acting toxic bait. However, the ability for S. marcescens and P. aeruginosa to survive on wood is low, which made the bait unable to retain a useful level of toxicity for a long period of time and frequent reapplication is needed.
Some novel hydrazone derivatives 6a-o were synthesized from the key intermediate 4-Chloro-N-(2-hydrazinocarbonyl-phenyl)-benzamide 5 and characterized using IR, ¹H-NMR, 13C-NMR, mass spectroscopy and elemental analysis. The inhibitory potential against two secretory phospholipase A₂ (sPLA₂), three protease enzymes and eleven bacterial strains were evaluated. The results revealed that all compounds showed preferential inhibition towards hGIIA isoform of sPLA₂ rather than DrG-IB with compounds 6l and 6e being the most active. The tested compounds exhibited excellent antiprotease activity against proteinase K and protease from Bacillus sp. with compound 6l being the most active against both enzymes. Furthermore, the maximum zones of inhibition against bacterial growth were exhibited by compounds; 6a, 6m, and 6o against P. aeruginosa; 6a, 6b, 6d, 6f, 6l, 6m, 6n, and 6o against Serratia; 6k against S. mutans; and compounds 6a, 6d, 6e, 6m, and 6n against E. feacalis. The docking simulations of hydrazones 6a-o with GIIA sPLA₂, proteinase K and hydrazones 6a-e with glutamine-fructose-6-phosphate transaminase were performed to obtain information regarding the mechanism of action.
Serratia marcescens is an opportunistic bacterial pathogen with broad range of host ranging from vertebrates, invertebrates and plants. S. marcescens strain W2.3 was isolated from a diseased tilapia fish and it was suspected to be the causal agent for the fish disease as virulence genes were found within its genome. In this study, for the first time, the genome sequences of S. marcescens strain W2.3 were sequenced using the Illumina MiSeq platform.
Here, we present the first complete genome sequence of Serratia fonticola DSM 4576(T), a potential plant growth promoting (PGP) bacterium which confers solubilization of inorganic phosphate, indole-3-acetic acid production, hydrogen cyanideproduction, siderophore production and assimilation of ammonia through the glutamate synthase (GS/GOGAT) pathway. This genome sequence is valuable for functional genomics and ecological studies which are related to PGP and biocontrol activities.
Serratia multitudinisentens RB-25(T) (=DSM 28811(T) =LMG 28304(T)) is a newly proposed type strain in the genus of Serratia isolated from a municipal landfill site. Here, we present the complete genome of S. multitudinisentens RB-25(T) which contains a complete chitinase operon and other chitin and N-acetylglucosamine utilisation enzymes. To our knowledge, this is the first report of the complete genome sequence of this novel isolate and its chitinase gene discovery.
Quorum sensing is a unique bacterial communication system which permits bacteria to synchronize their behaviour in accordance with the population density. The operation of this communication network involves the use of diffusible autoinducer molecules, termed N-acylhomoserine lactones (AHLs). Serratia spp. are well known for their use of quorum sensing to regulate the expression of various genes. In this study, we aimed to characterized the AHL production of a bacterium designated as strain RB-25 isolated from a former domestic waste landfill site. It was identified as Serratia fonticola using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry analysis and this was confirmed by 16S ribosomal DNA sequencing. High resolution triple quadrupole liquid chromatography-mass spectrometry analysis of S. fonticola strain RB-25 spent culture supernatant indicated the existence of three AHLs namely: N-butyryl-L-homoserine lactone (C4-HSL), N-hexanoyl-L-homoserine lactone (C6-HSL) and N-(3-oxohexanoyl) homoserine-lactone (3-oxo-C6 HSL). This is the first report of the production of these AHLs in S. fonticola.
Burkholderia cenocepacia and Serratia marcescens are Gram-negative proteobacteria commonly found in the natural
environment and are also opportunistic pathogens that caused a number of human diseases. The fermentation culture of
Burkholderia cenocepacia yielded three compounds, 4-(2-hydroxyethoxy)-phenol (1), Maculosin (2) and methyl myristate
(3). Compound 2 was also isolated together with cyclo(L-Leu-L-Pro) (4) from Serratia marcescens. Compound 1 was
isolated from a natural source for the first time and the first isolation of compounds 2-4 was also reported from both
Burkholderia cenocepacia and Serratia marcescens.
The σ(S) subunit RpoS of RNA polymerase functions as a master regulator of the general stress response in Escherichia coli and related bacteria. RpoS has been reported to modulate biocontrol properties in the rhizobacterium Serratia plymuthica IC1270. However, the role of RpoS in the stress response and biofilm formation in S. plymuthica remains largely unknown. Here we studied the role of RpoS from an endophytic S. plymuthica G3 in regulating these phenotypes. Mutational analysis demonstrated that RpoS positively regulates the global stress response to acid or alkaline stresses, oxidative stress, hyperosmolarity, heat shock and carbon starvation, in addition to proteolytic and chitinolytic activities. Interestingly, rpoS mutations resulted in significantly enhanced swimming motility, biofilm formation and production of the plant auxin indole-3-acetic acid (IAA), which may contribute to competitive colonization and environmental fitness for survival. These findings provide further insight into the strain-specific role of RpoS in the endophytic strain G3 of S. plymuthica, where it confers resistance to general stresses encountered within the plant environment. The heterogeneous functionality of RpoS in rhizosphere and endophytic S. plymuthica populations may provide a selective advantage for better adaptation to various physiological and environmental stresses.
The antibiotic pyrrolnitrin (PRN) is a tryptophan-derived secondary metabolite that plays an important role in the biocontrol of plant diseases due to its broad-spectrum of antimicrobial activities. The PRN biosynthetic gene cluster remains to be characterised in Serratia plymuthica, though it is highly conserved in PRN-producing bacteria. To better understand PRN biosynthesis and its regulation in Serratia, the prnABCD operon from S. plymuthica G3 was cloned, sequenced and expressed in Escherichia coli DH5α. Furthermore, an engineered strain prnind which is a conditional mutant of G3 prnABCD under the control of the Ptac promoter was constructed. This mutant was able to overproduce PRN with isopropylthiogalactoside (IPTG) induction by overexpressing prnABCD, whilst behaving as a conditional mutant of G3 prnABCD in the absence of IPTG. These results confirmed that prnABCD is responsible for PRN biosynthesis in strain G3. Further experiments involving lux-/dsRed-based promoter fusions, combined with site-directed mutagenesis of the putative σS extended -10 region in the prnA promoter, and liquid chromatography-mass spectrometry (LC-MS) analysis extended our previous knowledge about G3, revealing that quorum sensing (QS) regulates PRN biosynthesis through cross talk with RpoS, which may directly activated prnABCD transcription. These findings suggest that PRN in S. plymuthica G3 is produced in a tightly controlled manner, and has diverse functions, such as modulation of cell motility, in addition to antimicrobial activities. Meanwhile, the construction of inducible mutants could be a powerful tool to improve PRN production, beyond its potential use for the investigation of the biological function of PRN.
Molybdenum, an emerging pollutant, has being demonstrated recently to be toxic to
spermatogenesis in several animal model systems. Metal mines especially gold mine often use
cyanide and hence isolation of metal-reducing and cyanide-degrading bacteria can be useful for
the bioremediation of these pollutants. Preliminary screening shows that three cyanide-degrading
bacteria were able to reduce molybdenum to molybdenum blue (Mo-blue) when grown on a
molybdate low phosphate minimal salts media. Phylogenetic analyses of the 16S rRNA gene of
the best reducer indicates that it belongs to the Serratia genus. A variety of mathematical models
such as logistic, Gompertz, Richards, Schnute, Baranyi-Roberts, von Bertalanffy, Buchanan
three-phase and Huang were used to model molybdenum reduction, and the best model based on
statistical analysis was modified Gompertz with lowest values for RMSE and AICc, highest
adjusted R2 values, with Bias Factor and Accuracy Factor nearest to unity (1.0). The reduction
constants obtained from the model will be used to carry out secondary modelling to study the
effect of various parameters such as substrate, pH and temperature to molybdenum reduction.
The volume of contaminated rivers in Malaysia continues to keep rising through the years. The
cost of instrumental monitoring is uneconomical and prohibits schedule monitoring of
contaminants particularly heavy metals. In this work, a rapid enzyme assay utilizing the
molybdenum-reducing enzyme as an inhibitive assay, prepared in crude form from the
molybdenum-reducing bacterium Serratia sp. strain DRY5 has been developed for monitoring
the heavy metals mercury, silver, copper and chromium in contaminated waters in the Juru
Industrial Estate. The crude enzyme extract transformed soluble molybdenum
(phosphomolybdate) into a deep blue solution, which is inhibited by heavy metals such as
mercury, silver, copper and chromium. The IC50 and Limits of Detection (LOD) values for
mercury, copper, silver and cadmium were 0.245, 0.298, 0.367, 0.326, and 0.124, 0.086, 0.088
and 0.094 mg L-1, respectively. The assay is rapid, and can be carried out in less than 10 minutes.
In addition, the assay can be carried out at ambient temperature. The IC50 values for these heavy
metals are more sensitive than several established assays. Water samples from various locations
in the month of November from the Juru Industrial Estate (Penang) were tested for the presence
of heavy metals using the developed assay. Enzyme activity was nearly inhibited for water
samples from several locations. The presence of heavy metals was confirmed instrumentally
using Atomic Emission Spectrometry and a Flow Injection Mercury System. The assay is rapid
and simple and can be used as a first screening method for large scale monitoring of heavy
metals.
Serratia marcescens subsp.sakuensisstrain K27 was isolated from sponge (Haliclona amboinensis). The genome of this strain consists of 5,325,727 bp, with 5,140 open reading frames (ORFs), 3 rRNAs, and 67 tRNAs. It contains genes for the production of amylases, lipases, and proteases. Gene clusters for the biosynthesis of nonribosomal peptides and thiopeptide were also identified.
For the past few decades, there has been limited progress in the development of novel antibacterials. Previously, we postulated that the gut microbiota of animals residing in polluted environments are a forthcoming supply of antibacterials. Among various species, the water monitor lizard is an interesting species that feeds on organic waste and the carcass of wild animals. Gut microbiota of the water monitor lizard were sequestered, identified and cultivated in RPMI-1640 to produce conditioned medium (CM). Next, the antimicrobial properties of CM were evaluated versus a selection of Gram-negative (Escherichia coli K1, Serratia marcescens,Pseudomonas aeruginosa, Salmonella enterica and Klebsiella pneumoniae) and Gram-positive bacteria (Streptococcus pyogenes, methicillin-resistant Staphylococcus aureus, and Bacillus cereus). CM were partially characterized by heat inactivation at 95°C for 10 min and tested against P. aeruginosa and S. pyogenes. CM were also tested against immortalized human keratinocytes (HaCaT) cells lines. The results demonstrated that gut microbiota isolated from water monitor lizard produced molecules with remarkable bactericidal activities. To determine the identity of the active molecules, CM were subjected to Liquid Chromatography-Mass Spectrometry. Several molecules were identified belonging to the classes of flavonoids, terpenoids, alkaloids, polyhydroxy alkaloids, polyacetylenes, bisphenols, amides, oxylipin and pyrazine derivatives with known broad-spectrum antimicrobial, anti-tumour, anti-oxidant, anti-inflammatory, and analgesic attributes. Furthermore, the detailed analysis of these molecules could lead us to develop effective therapeutic antibacterials.
Among several animals, Rattus rattus (rat) lives in polluted environments and feeds on organic waste/small invertebrates, suggesting the presence of inherent mechanisms to thwart infections. In this study, we isolated gut bacteria of rats for their antibacterial activities. Using antibacterial assays, the findings showed that the conditioned media from selected bacteria exhibited bactericidal activities against Gram-negative (Escherichia coli K1, Klebsiella pneumoniae, Pseudomonas aeruginosa, Serratia marcescens, and Salmonella enterica) and Gram-positive (Bacillus cereus, methicillin-resistant Staphylococcus aureus, and Streptococcus pyogenes) pathogenic bacteria. The conditioned media retained their antibacterial properties upon heat treatment at boiling temperature for 10 min. Using MTT assays, the conditioned media showed minimal cytotoxic effects against human keratinocyte cells. Active conditioned media were subjected to tandem mass spectrometry, and the results showed that conditioned media from Bacillus subtilis produced a large repertoire of surfactin and iturin A (lipopeptides) molecules. To our knowledge, this is the first report of isolation of lipopeptides from bacteria isolated from the rat gut. In short, these findings are important and provide a platform to develop effective antibacterial drugs.
Type A chitinases (EC 3.2.1.14), GH family 18, attack chitin ((1 → 4)-2-acetamido-2-deoxy-β-D-glucan) and chito-oligosaccharides from the reducing end to catalyze release of chitobiose (N,N'-diacetylchitobiose) via hydrolytic cleavage of N-acetyl-β-D-glucosaminide (1 → 4)-β-linkages and are thus "exo-chitobiose hydrolases." In this study, the chitinase type A from Serratia marcescens (SmaChiA) was used as a template for identifying two novel exo-chitobiose hydrolase type A enzymes, FbalChi18A and MvarChi18A, originating from the marine organisms Ferrimonas balearica and Microbulbifer variabilis, respectively. Both FbalChi18A and MvarChi18A were recombinantly expressed in Escherichia coli and were confirmed to exert exo-chitobiose hydrolase activity on chito-oligosaccharides, but differed in temperature and pH activity response profiles. Amino acid sequence comparison of the catalytic β/α barrel domain of each of the new enzymes showed individual differences, but ~69% identity of each to that of SmaChiA and highly conserved active site residues. Superposition of a model substrate on 3D structural models of the catalytic domain of the enzymes corroborated exo-chitobiose hydrolase type A activity for FbalChi18A and MvarChi18A, i.e., substrate attack from the reducing end. A main feature of both of the new enzymes was the presence of C-terminal 5/12 type carbohydrate-binding modules (SmaChiA has no C-terminal carbohydrate binding module). These new enzymes may be useful tools for utilization of chitin as an N-acetylglucosamine donor substrate via chitobiose.
Introduction: The aim of this study is to determine the most common organisms isolated in diabetic foot infection and the most utilised antibiotic regimes as the first line of treatment.
Methods: This is a retrospective record review of the National Orthopaedic Registry Malaysia among diabetes mellitus type 2 patients who had foot infections. All identified cases admitted to 18 government hospitals in Malaysia from the 1st January 2008 until the 31st December, 2009 were included in the study.
Results: A total of 416 patients were included in the study. The most common organisms cultured were Proteus species (17.5%), Klebsiella species (17.1%) and Staphylococcus aureus (17.9%), while the most commonly used antibiotic was ampicillin/sulbactam (67.5%). None of the patients was appropriately treated with metronidazole, cefoperazone or fucidic acid. All patients were given appropriate antibiotics to treat Serratia infection.
Conclusion: Significant number of patients with diabetic foot infections were not treated using appropriate antibiotics as the first line treatment.