Bacteriocin Activity Research Articles

Heterologous expression and purification of BtCspB, a novel cold-shock protein-like bacteriocin from Bacillus thuringiensis BRC-ZYR2.

A novel Bacillus thuringiensis (Bt) bacteriocin BtCspB, active against a food-borne pathogen Bacillus cereus, was identified and purified by a traditional four-step chromatographic process with low yield (44.5µg/L) in our lab previously. The aim of this study was to dramatically increase its yield by heterologous expression of BtCspB. The BtCspB gene from Bt BRC-ZYR2 was successfully heterologously expressed in Escherichia coli BL21 (DE3). Affinity chromatography was used to obtain the pure BtCspB up to 20mg/L. The purified BtCspB showed a MIC value of 12.5µg/mL and a MBC value of 50.0µg/mL against Bacillus cereus ATCC 10987. The bacteriocin activity of BtCspB against B. cereus ATCC 10987 was further directly detected in a gel-overlay assay. The anti-B. cereus activity, however, was lower than the bacteriocin purified by the traditional four-step chromatographic process probably because of structural modifications. Compared with the traditional method, the yield of the bacteriocin by heterologous expression increased by 449 times, and the purification step was dramatically simplified, which laying a foundation for the industrial production of this novel cold-shock protein-like bacteriocin BtCspB active against B. cereus.

Preservation of Meat Products with Bacteriocins Produced by Lactic Acid Bacteria Isolated from Meat

Bacteriocins are ribosomal-synthesized antimicrobial peptides that inhibit the growing of pathogenic and/or deteriorating bacteria. The most studied bacteriocin-producing microorganisms are lactic acid bacteria (LAB), as they have great potential application in food biopreservation, since the majority have GRAS (Generally Recognized as Safe) status. The LAB-producing bacteriocins and/or bacteriocins produced by these bacteria have been widely studied, with the emphasis on those derived from milk and dairy products. On the other hand, isolates from meat and meat products are less studied. The objective of this review is to address the main characteristics, classification, and mechanism of action of bacteriocins and their use in food, to highlight studies on the isolation of LAB with bacteriocinogenic potential from meat and meat products and also to characterize, purify, and apply these bacteriocins in meat products. In summary, most of the microorganisms studied areLactococcus,Enterococcus,Pediococcus, andLactobacillus, which produce bacteriocins such as nisin, enterocin, pediocin, pentocin, and sakacin, many with the potential for use in food biopreservation.

Antimicrobial efficacy of the supernatant of Streptococcus dentisani against microorganisms implicated in root canal infections.

The present study aimed to test the antimicrobial activity of Streptococcus dentisani (S. dentisani) supernatant against a collection of microorganisms implicated in dental root infections, and to analyze morphological changes induced in a selection of the tested microorganisms. A total of 22 microbial species were selected, and their growth was monitored by spectrophotometry in the presence and absence of the supernatant of S. dentisani at different assay concentrations (0.2×, 1×, 2×). The generation time and maximum growth rates were evaluated under every tested condition. Scanning electron microscope (SEM) images were obtained to assess the effect on the cell surface following incubation of the pathogens with the concentrated (2×) supernatant of S. dentisani. The supernatant of S. dentisani was found to exert effective inhibitory activity against most of the studied microorganisms implicated in dental root infections (20 out of 22). Total growth inhibition was observed in the case of Streptococcus oralis, Streptococcus sobrinus, Streptococcus salivarius, Prevotella intermedia, and Streptococcus mutans, while the rest of the microorganisms showed an increase in the generation time (between 30 min and 4 h). SEM images revealed structural changes in the membrane consistent with bacteriocin activity, although the effects were heterogeneous among the different species tested.

Anti-staphylococcal activity of bacteriocins of food isolates Enterococcus hirae LD3 and Lactobacillus plantarum LD4 in pasteurized milk.

Bacteriocins of Enterococcus hirae LD3 and Lactobacillus plantarum LD4 have been applied in milk for growth inhibition of Staphylococcus aureus. The enumeration of S. aureus cells in nutrient broth and milk was found log10 9.7 and 10.2CFU/mL, respectively, whereas it was reduced with increasing concentration of bacteriocins suggesting loss of cell viability. The lethal concentration (LC50) of enterocin LD3 and plantaricin LD4 against S. aureus was 160 and 220µg/mL, respectively. Bacteriocin-treated cells were stained red with propidium iodide (PI) indicating dead cells further confirms bactericidal nature. The enterocin LD3-treated cells showed higher infrared absorbance at 1451.82cm- 1 corresponding to phospholipids suggesting membrane-acting nature of the bacteriocin. However, plantaricin LD4-treated cells did not show such alterations suggesting different mode of action. Both bacteriocins caused disruption and shrinkage of target cells, and leakage of intracellular contents as observed in transmission electron microscope (TEM). The present study suggests killing of S. aureus in milk, therefore, enterocin LD3 and plantaricin LD4 may be applied in biopreservation of milk and related food products.

Antimicrobial activity of enterocin obtained from Enterococcus durans on Shiga-like toxin-producing Escherichia coli

ABSTRACT: Shiga-like toxin-producing Escherichia coli (STEC) is an important source of food contamination that presents risks to human health. Several industrial food processes eliminate this microorganism; however, these processes can alter the characteristics of the product. Alternative methods of preservation have been identified as an option to control these foodborne pathogens. The purpose of this study was to evaluate the action of bacteriocins produced by Enterococcus durans MF5 in STEC cells. Cell-free supernatant (CFS) containing enterocins from the MF5 isolate was tested over different time points (6, 18, and 24 h). Enterocins present in the crude CFS showed inhibition against STEC at all time points. In the investigation of cell integrity, using propidium iodide and fluorescence microscopy, considerable cell death was observed within 6 h of the cells being in contact with the enterocins, which was also observed at the 18 and 24 h time points. These results showed that the enterocins produced by the MF5 isolate have potential use in the control of STEC.

Antibacterial Activity Test of Bacteriocin from Lactobacillus brevis, Lactobacillus casei and Lactobacillus plantarum Against Gram Positive Pathogenic Bacteria

Bacteriocin is a secondary metabolite product of lactic acid bacteria (LAB) which have an antimicrobial and potentially as a natural preservative. LAB isolates used in this study were Lactobacillus brevis, Lactobacillus casei and Lactobacillus plantarum. This study aimed to determine the antibacterial activity of bacteriocin produced by each isolate of LAB including the influence of pH and heating variation against Bacillus cereus, Bacillus subtilis and Staphylococcus epidermidis. Antibacterial activity test was done by using disc diffusion method. method. Confirmation test using proteolytic enzyme aimed to analyse that the inhibition zone produced from the activity of bacteriocin. The inhibition zone produced from L. brevis, L. casei and L. plantarum against B. cereus were 15.70, 16.43 and 14.50 mm, against B. subtilis were 13.37, 14.10 and 12.53 mm and against S. epidermidis were 11.37, 14.50 and 12.45 mm. The activity of each bacteriocin decreased with the addition of trypsin and catalase, bacteriocin was active in the pH range of 2-10 and heating temperature of 40-121oC. Statistical test showed that the addition of trypsin, catalase and the variation of pH also heating had significant differences (p<0.05) to antibacterial activity produced by bacteriocin from L. brevis, L. casei and L. plantarum.

Antibacterial activity of crud Bacteriocin- like substance against food borne bacterial pathogens

Lactobacillus is one of Lactic Acid Bacteria group, they are known to exhibit antagonistic activity against pathogenic organisms. This study evaluates the protective effect of Lactobacillus acidophilus Crud Bacteriocin like substance against growth of some food borne pathogenic bacteria. Antimicrobial effect of crud bacteriocin like substance was tested against the growth of S. typhi and S. aureus that isolated from food samples by agar well diffusion assay.The results showed that the crud extract of bacteriocin like substance had antibacterial activity against the two tested bacteria, and the effect against S. typhi was greater than that of S. aureus , the zone of inhibition was (15)mm against S. typhi and (12)mm against S. aureus . The antibiotic susceptibility test was also studied against the two foodborne pathogenic bacteria.This study aims to screen the antibacterial activity of crud bacteriocin like substance from lactic acid bacteria against two food borne pathogens.

Intracellular pHluorin as Sensor for Easy Assessment of Bacteriocin-Induced Membrane-Damage in Listeria monocytogenes.

Bacteriocins are antimicrobial peptides naturally produced by many bacteria and were shown to be effective against various pathogens including Listeria monocytogenes. L. monocytogenes is a food-borne pathogen that frequently causes disease outbreaks around the world with fatal outcomes in at-risk individuals. Thus, bacteriocins are a promising solution to prevent contaminations with L. monocytogenes and other microorganisms during food production and preservation. In the present study, we constructed L. monocytogenes EGD-e/pNZ-Phelp-pHluorin, a strain that constitutively expresses the pH-sensitive fluorescent protein pHluorin, as a sensor strain to detect disruption of the pH gradient by the membrane-damaging activity of bacteriocins. The ratiometric fluorescence properties of pHluorin were validated both in crude extracts and permeabilized cells of this sensor strain. L. monocytogenes EGD-e/pNZ-Phelp-pHluorin was used to assess membrane damaging activity of the bacteriocins nisin A and pediocin PA-1 and to determine the minimal concentrations required for full disruption of the pH gradient across the membrane. Moreover, the sensor strain proved useful to analyze the presence of compounds affecting membrane integrity in supernatants of a nisin Z-producing Lactococcus lactis strain at different timepoints during growth. Supernatants of this strain that were active in disrupting the pH gradient across the membrane were also shown to inhibit growth of L. monocytogenes. In summary, the presented results suggest that the generated sensor strain is a convenient, fast and reliable tool to identify and characterize novel bacteriocins and other compounds that target membrane integrity.

Characterization and safety evaluation of partially purified bacteriocin produced by Escherichia coli E isolated from fermented pineapple Ananas comosus (L.) Merr.

Antibacterial activity of cell-free supernatant from Escherichia coli E against selected pathogenic bacteria in food and aquaculture was the highest against Edwardsiella tarda 3, a significant aquaculture pathogen. Biochemical properties of the bacteriocins were studied and bacteriocin was found to be sensitive to proteinase K, demonstrating its proteinaceous nature. In addition, pH and temperature affected bacteriocin activity and stability. The bacteriocins were partially purified by ammonium sulfate precipitation. The antibacterial activity was only detected in 20% ammonium sulfate fraction and direct detection of its activity was performed by overlaying on the indicator strains. The inhibition zone associated with the antibacterial activity was detected in the sample overlaid by E. tarda 3 and Staphylococcus aureus DMST8840 with the relative molecular mass of about 27kDa and 10kDa, respectively. Bacteriocin showed no cytotoxic effect on NIH-3T3 cell line; however, two virulence genes, aer and sfa, were detected in the genome of E. coli E by PCR. The characteristics of bacteriocins produced by E. coli E exhibited the antibacterial activity against both Gram-positive and Gram-negative pathogenic bacteria and the safe use determined by cytotoxicity test which may have interesting biotechnological applications.

Enhanced Bacteriocin Production by Pediococcus pentosaceus 147 in Co-culture With Lactobacillus plantarum LE27 on Cheese Whey Broth.

The production of bacteriocins by lactic acid bacteria (LAB) has been of wide interest in the food industry due to their potential application in biopreservation. The production of bacteriocins is usually low in single strain fermentation, but can improve when the bacteriocinogenic strain is cultured in association with another bacteria. The present work aims to evaluate the growth and production of bacteriocins by Pediococcus pentosaceus 147 (bacteriocinogenic strain) in co-culture with Lactobacillus plantarum LE27 (inducer strain) using a culture medium based on cheese whey (CW). Strains were inoculated in co-culture in a CW broth at 7.24 Log CFU/mL of initial concentration of P. pentosaceus 147 and incubated at 37°C. Bacteriocin production was measured after 24 h by the critical dilution method, biomass was measured by plating on MRS agar (1% aniline blue), and a mono-culture was used as a control. The titers of bacteriocins produced by P. pentosaceus 147 in mono-culture were 19,200 AU/mL lower than those obtained in co-culture with Lb. plantarum LE27 at 51,200 AU/mL. The effect of adding the inducer strain at different times of incubation (3, 6, 9, and 12 h) was evaluated, with the addition of the induction factor at the beginning of the incubation of P. pentosaceus 147 generating the highest bacteriocin activity. This study shows the potential of inducing bacteriocinogenesis using co-cultures of strains of the genera Pediococcus and Lactobacillus and using alternative substrates such as cheese whey.

REDUCTION OF PATHOGENIC BACTERIA DURING FERMENTATION OF MASIN BY PROTEASE AND BACTERIOCIN-PRODUCING LACTIC ACID BACTERIA

Contamination of pathogenic and spoilage bacteria on fermentation process of seafood and fisheries product is a major concern on food safety. The aims of this study were isolating and applying the bacteriocin- and protease-producing lactic acid bacteria from the gastrointestinal tract of mud crab for starter culture of masin, a traditional fermented shrimp from East Java. This study consisted of characterization of lactic acid bacteria, the application on the fermentation process and microbial analysis. Ninety-four isolates were isolated from mud crab was screened for the bacteriocin and protease producing as well as characterized by pH, salinity and biochemical. Isolate IKP-29 was exhibited strong protease and bacteriocin activity. Application of Isolate IKP-29 on masin fermentation showed that sharp reduction of Escherichia coli, Vibrio sp. and lactic acid bacteria counted. This study suggested that the application of lactic acid bacteria which producing bacteriocin and protease improved the food safety of traditional fermented fish.

In-Vitro Study the Antibacterial Activity of Bacteriocin against Stenotrophomonas Maltophilia and Evaluation its Synergism with some Antibiotics

In-Vitro Study the Antibacterial Activity of Bacteriocin against Stenotrophomonas Maltophilia and Evaluation its Synergism with some Antibiotics

New Approach in Acne Therapy: A Specific Bacteriocin Activity and a Targeted Anti IL-8 Property in Just 1 Probiotic Strain, the L. salivarius LS03.

The aim of this research was to assess the antibacterial activity of Lactobacillus salivarius LS03 (DSM 22776) against Propionibacterium acnes and its anti-inflammatory properties by inhibiting P. acnes-induced interleukin-8 (IL-8) release. Acne is the most common skin disease, causing significant psychosocial problems for those afflicted. Currently available agents for acne treatment, such as oral antibiotics, have limited use. Thus, development of novel agents to treat this disease is needed. In the generation of inflammatory lesions, proliferation of P. acnes in the obstructed follicles is critical. The administration of beneficial microorganisms represents a promising approach for treating several skin alterations and can have many favorable effects. For the inhibition assay, P. acnes was spread on Propionibacter Isolation Agar Base plates, and LS03-soaked disks were placed directly on the agar surface. Peripheral blood mononuclear cells, isolated from healthy volunteers, were preincubated with phytohemagglutinin 1 μg/mL for 1 hour and stimulated with the probiotic strains for 24 hours to simulate an in vitro IL-8 release model. The IL-8 concentration in the supernatants was analyzed in duplicate using ELISA Kit. L. salivarius LS03 exerted a significant inhibitory capacity against the target pathogen strain. This antagonistic activity was primarily ascribable to the feature of LS03 strain of secreting active bacteriocins against P. acnes. Concerning the IL-8 analysis, 3 different L. salivarius strains were able to inhibit the release of this chemokine by 10% to 25%. L. salivarius LS03 probiotic strain could be an alternative treatment to antibiotic/anti-inflammatory therapy in subjects presenting acne vulgaris.

Antibiofilm activity of klebocin crude extract against some species of Enterobacteriaceae

Bacteriocins are antibacterial proteins created by bacteria and its effective against other strains of bacteria which are closely linked to the producing strains and a number of species from the same family. The aim of current study was to evaluate the activity of bacteriocin that extracted from clinical isolates of Klebsiella pneumonia against different species of enterobacteriaceae in both planktonic and biofilm state. The antibacterial activity of bacteriocins (klebocins) from Klebsiella pneumoniae isolates towards diverse pathogenic species from enterobacteriaceae (by well assay method) was demonstrated , however the antibiofilm activity against some species of enterobacteriaceae was studied also by Tissue culture plate method (TCP) in first and last stages of biofilm formation .The results revealed that klebocin with different concentrations was efficient against different pathogenic species by producing different inhibition zones. Results also showed that klebocins had a wide antibiofilm activity on some pathogenic species of Gram-negative bacteria .In addition to these finding it was observed that the antibiofilm activity of klebocin on premature biofilm of Klebsiella was higher than its effect towards other pathogenic species, while it was affected on mature biofilm of other bacterial species as well as Klebsiella.

Antimicrobial Activity of a Bacteriocin Produced by Enterococcus faecalis KT11 against Some Pathogens and Antibiotic-Resistant Bacteria.

In this study, the antimicrobial activity of a bacteriocin produced by Enterococcus faecalis KT11, isolated from traditional Kargı Tulum cheese, was determined, and bacteriocin KT11 was partially characterized. The results showed that bacteriocin KT11 was antagonistically effective against various Gram-positive and Gram-negative test bacteria, including vancomycin- and/or methicillin-resistant bacteria. The activity of bacteriocin KT11 was completely abolished after treatment with proteolytic enzymes (proteinase K, α-chymotrypsin, protease and trypsin), which demonstrates the proteinaceous nature of this bacteriocin. Additionally, bacteriocin KT11 remained stable at pH values ranging from 2 to 11 and after autoclaving at 121℃ for 30 min. In addition, the activity of bacteriocin KT11 was stable after treatment with several surfactants (EDTA, SDS, Triton X-100, Tween 80 and urea) and organic solvents (chloroform, propanol, methanol, ethyl alcohol, acetone, hexane and ethyl ether). Cell-free supernatant of E. faecalis KT11 was subjected to ammonium sulfate precipitation and then desalted by using a 3.5-kDa cut-off dialysis membrane. The bacteriocin activity was determined to be 711 AU/mL in the dialysate. After tricine-SDS-PAGE analysis, one peptide band, which had a molecular weight of ~3.5 kDa, exhibited antimicrobial activity. Because the bacteriocin KT11, isolated from E. faecalis KT11, exhibits a broad antimicrobial spectrum, heat stability and stability over a wide pH range, this bacteriocin can be used as a potential bio-preservative in foods. Additionally, bacteriocin KT11 alone or in combination with conventional antibiotics may provide a therapeutic option for the treatment of multidrug-resistant clinical pathogens after further in vivo studies.

Purification and Partial Characterization of Bacteriocin Lac-B23, a Novel Bacteriocin Production by Lactobacillus plantarum J23, Isolated From Chinese Traditional Fermented Milk.

The exploration and evaluation of bacteriocin-producing lactic acid bacteria (LAB) have been one of the powerful means to food preservation. A total of 300 strains were isolated from Chinese traditional fermented milk products. A bacteriocin-producing LAB, named Lactobacillus plantarum J23, was screened and identified. Bacteriocin Lac-B23 from L. plantarum J23 was purified by 80% ammonium sulfate precipitation, cation-exchange chromatography, and reverse-phase high-performance liquid chromatography. Molecular weight of bacteriocin Lac-B23 was determined to be approximately 6.73 kDa by tricine sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis, and it was confirmed as a novel bacteriocin by liquid chromatography-mass spectrometry. Moreover, bacteriocin Lac-B23 showed thermal stability when heated at below 100°C for 30 min, pH stability between pH 2.0 and 12.0, and sensitivity to trypsin, proteinase K, and proteinase E. The antimicrobial activity of bacteriocin could be enhanced by addition of Fe2+, Mn2+, and ethyl alcohol, and inhibited by Cu2+, K+, Ca2+, Zn2+, Mg2+, and sodium chloride. The results suggested bacteriocin Lac-B23 to have potential application prospects in the food industry.

Bacteriocin production of the probiotic Lactobacillus acidophilus KS400

In the last years, the use of probiotics, including Lactobacillus species, has received much attention to prevent and treat vaginal disorders. These species have been described as having the ability to colonize the epithelial surface and produce antimicrobial metabolites that are able to control the remaining vaginal microflora. This study aimed to identify and characterize, for the first time, a bacteriocin natively produced by Lactobacillus acidophilus KS400 (probiotic strain from Gynoflor®-Medinova AG, Switzerland) and its antimicrobial activity against relevant urogenital pathogens. After organic acids and hydrogen peroxide neutralization in the fermented Lactobacillus acidophilus KS400 culture medium, bacteriocin activity was tested against the indicator microorganism Lactobacillus delbrueckii ATCC9649. The fermentation of Lactobacillus acidophilus KS400 for bacteriocin production was carried out in batch mode, and its antimicrobial activity, optical density and pH were monitored. After production and extraction, the bacteriocin molecular weight was estimated by electrophoresis and tested against vaginal pathogenic microorganisms. As described for other bacteriocins, batch fermentation profiles indicated that bacteriocin production occurs during the exponential growth phase of the lactobacilli, and declines during their stationary growth phase. The molecular weight of the bacteriocin is approximately 7.5 kDa. The bacteriocin containing protein extract was shown to inhibit the growth of Gardnerella vaginalis, Streptococcus agalactiae, Pseudomonas aeruginosa and the indicator strain Lactobacillus delbrueckii ATCC9649. We conclude that L. acidophilus KS400 produces bacteriocin with antimicrobial activity against relevant urogenital pathogens.

Mode of action of a novel anti-Listeria bacteriocin (CAMT2) produced by Bacillus amyloliquefaciens ZJHD3-06 from Epinephelus areolatus.

Bacteriocin CAMT2, produced by Bacillus amyloliquefaciens ZJHD3-06, has been shown to exhibit protective activity against important food spoilage and food-borne bacterial pathogens. This study was conducted to investigate the mode of action of bacteriocin CAMT2 against highly pathogenic Listeria monocytogenes ATCC 19111. The addition of bacteriocin CAMT2 at 64AU/ml inhibited L. monocytogenes ATCC 19111. An efflux of K+ ions, lactic acid dehydrogenase and an increase in extracellular electrical conductivity was observed in CAMT2-treated L. monocytogenes. Electron microscopy showed morphological alterations such as uneven cell surface, accumulation of cell debris and bacterial lysis. These results show that bacteriocin CAMT2 inhibit L. monocytogenes by increasing cell permeability and inducing membrane damage, hence it has the great application potentials in ensuring food safety.

Localized recombination drives diversification of killing spectra for phage-derived syringacins

To better understand the potential for antagonistic interactions between members of the same bacterial species, we have surveyed bacteriocin killing activity across a diverse suite of strains of the phytopathogen Pseudomonas syringae. Our data demonstrate that killing activity from phage-derived bacteriocins of P. syringae (R-type syringacins) is widespread. Despite a high overall diversity of bacteriocin activity, strains can broadly be classified into five main killing types and two main sensitivity types. Furthermore, we show that killing activity switches frequently between strains and that switches correlate with localized recombination of two genes that together encode the proteins that specify bacteriocin targeting. Lastly, we demonstrate that phage-derived bacteriocin killing activity can be swapped between strains simply through expression of these two genes in trans. Overall, our study characterizes extensive diversity of killing activity for phage-derived bacteriocins of P. syringae across strains and highlights the power of localized recombination to alter phenotypes that mediate strain interactions during evolution of natural populations and communities.

A Colicin M-Type Bacteriocin from Pseudomonas aeruginosa Targeting the HxuC Heme Receptor Requires a Novel Immunity Partner.

Pyocins are bacteriocins secreted by Pseudomonas aeruginosa, and they assist in the colonization of different niches. A major subset of these antibacterial proteins adopt a modular organization characteristic of polymorphic toxins. They include a receptor-binding domain, a segment enabling membrane passage, and a toxin module at the carboxy terminus, which eventually kills the target cells. To protect themselves from their own products, bacteriocin-producing strains express an immunity gene concomitantly with the bacteriocin. We show here that a pyocin equipped with a phylogenetically distinct ColM toxin domain, PaeM4, mediates antagonism against a large set of P. aeruginosa isolates. Immunity to PaeM4 is provided by the inner membrane protein PmiC, which is equipped with a transmembrane topology not previously described for the ColM family. Given that strains lacking a pmiC gene are killed by PaeM4, the presence of such an immunity partner likely is a key criterion for escaping cellular death mediated by PaeM4. The presence of a TonB box in PaeM4 and enhanced bacteriocin activity under iron-poor conditions strongly suggested the targeting of a TonB-dependent receptor. Evaluation of PaeM4 activities against TonB-dependent receptor knockout mutants in P. aeruginosa PAO1 revealed that the heme receptor HxuC (PA1302) serves as a PaeM4 target at the cellular surface. Because other ColM-type pyocins may target the ferrichrome receptor FiuA, our results illustrate the versatility in target recognition conferred by the polymorphic nature of ColM-type bacteriocins.IMPORTANCE The antimicrobial armamentarium of a bacterium is a major asset for colonizing competitive environments. Bacteriocins comprise a subset of these compounds. Pyocins are an example of such antibacterial proteins produced by Pseudomonas aeruginosa, killing other P. aeruginosa strains. A large group of these molecules show a modular protein architecture that includes a receptor-binding domain for initial target cell attachment and a killer domain. In this study, we have shown that a novel modular pyocin (PaeM4) that kills target bacteria via interference with peptidoglycan assembly takes advantage of the HxuC heme receptor. Cells can protect themselves from killing by the presence of a dedicated immunity partner, an integral inner membrane protein that adopts a transmembrane topology distinct from that of proteins currently known to provide immunity against such toxin activity. Understanding the receptors with which pyocins interact and how immunity to pyocins is achieved is a pivotal step toward the rational design of bacteriocin cocktails for the treatment of P. aeruginosa infections.