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1.
Food Chem ; 449: 139229, 2024 Aug 15.
Article En | MEDLINE | ID: mdl-38581793

The unique high isoelectric point of lysozyme (LYZ) restricts its application in composite antibacterial coating due to the unfavorable liability to electrostatic interaction with other components. In this work, the antibacterial activity of a dispersible LYZ-carboxymethyl konjac glucomannan (CMKGM) polyelectrolyte complex was evaluated. Kinetic analysis revealed that, compared with free LYZ, the complexed enzyme exhibited decreased affinity (Km) but markedly increased Vmax against Micrococcus lysodeikticus, and QCM and dynamic light scattering analysis confirmed that the complex could bind with the substrate but in a much lower ratio. The complexation with CMKGM did not alter the antibacterial spectrum of LYZ, and the complex exerted antibacterial function by delaying the logarithmic growth phase and impairing the cell integrity of Staphylococcus aureus. Since the LYZ-CMKGM complex is dispersible in water and could be assembled easily, it has great potential as an edible coating in food preservation.


Anti-Bacterial Agents , Mannans , Muramidase , Staphylococcus aureus , Mannans/chemistry , Mannans/pharmacology , Mannans/metabolism , Muramidase/chemistry , Muramidase/metabolism , Muramidase/pharmacology , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/chemistry , Staphylococcus aureus/drug effects , Staphylococcus aureus/growth & development , Kinetics , Micrococcus/drug effects , Micrococcus/growth & development
2.
Biosci Biotechnol Biochem ; 88(5): 546-554, 2024 Apr 22.
Article En | MEDLINE | ID: mdl-38409797

Human lysozyme (hLYZ) has attracted considerable research attention due to its natural and efficient antibacterial abilities and widespread uses. In this study, hLYZ was modified to enhance its enzyme activity and expressed in a Pichia pastoris expression system. A combination mutant HZM(2R-K)-N88D/V110S demonstrated the highest enzyme activity (6213 ± 164 U/mL) in shake flasks, which was 4.07-fold higher when compared with the original strain. Moreover, the recombinant P. pastoris was inducted in a 3 L bioreactor plus methanol/sorbitol co-feeding. After 120 h induction, the antibacterial activity of hLYZ reached 2.23 ± 0.12 × 105 U/mL, with the specific activity increasing to 1.89 × 105 U/mg, which is currently the highest specific activity obtained through recombinant expression of hLYZ. Also, hLYZ supernatants showed 2-fold inhibitory effects toward Staphylococcus aureus and Micrococcus lysodeikticus when compared with HZM(2R-K). Our research generated a hLYZ mutant with high antibacterial capabilities and provided a method for screening of high-quality enzymes.


Anti-Bacterial Agents , Muramidase , Recombinant Proteins , Staphylococcus aureus , Muramidase/genetics , Muramidase/pharmacology , Muramidase/metabolism , Anti-Bacterial Agents/pharmacology , Humans , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Recombinant Proteins/pharmacology , Staphylococcus aureus/drug effects , Bioreactors , Micrococcus/drug effects , Gene Expression , Mutation , Saccharomycetales/genetics , Microbial Sensitivity Tests
3.
Appl Microbiol Biotechnol ; 108(1): 167, 2024 Jan 23.
Article En | MEDLINE | ID: mdl-38261012

As the problem of antimicrobial resistance is constantly increasing, there is a renewed interest in antimicrobial products derived from natural sources, particularly obtained from innovative and eco-friendly materials. Insect lipids, due to their fatty acid composition, can be classified as natural antimicrobial compounds. In order to assess the antibacterial efficacy of Hermetia illucens lipids, we extracted this component from the larval stage, fed on different substrates and we characterized it. Moreover, we analyzed the fatty acid composition of the feeding substrate, to determine if and how it could affect the antimicrobial activity of the lipid component. The antimicrobial activity was evaluated against Gram-positive Micrococcus flavus and Gram-negative bacteria Escherichia coli. Analyzing the fatty acid profiles of larval lipids that showed activity against the two bacterial strains, we detected significant differences for C4:0, C10:0, C16:1, C18:3 n3 (ALA), and C20:1. The strongest antimicrobial activity was verified against Micrococcus flavus by lipids extracted from larvae reared on strawberry, tangerine, and fresh manure substrates, with growth inhibition zones ranged from 1.38 to 1.51 mm, while only the rearing on manure showed the effect against Escherichia coli. Notably, the fatty acid profile of H. illucens seems to not be really influenced by the substrate fatty acid profile, except for C18:0 and C18:2 CIS n6 (LA). This implies that other factors, such as the rearing conditions, larval development stages, and other nutrients such as carbohydrates, affect the amount of fatty acids in insects. KEY POINTS: • Feeding substrates influence larval lipids and fatty acids (FA) • Generally, there is no direct correlation between substrate FAs and the same larvae FAs • Specific FAs influence more the antimicrobial effect of BSF lipids.


Diptera , Manure , Micrococcus , Animals , Larva , Escherichia coli , Fatty Acids , Micrococcus luteus
4.
R I Med J (2013) ; 107(2): 16-19, 2024 Feb 01.
Article En | MEDLINE | ID: mdl-38285745

Peritonitis, a serious complication of peritoneal dialysis (PD), can be caused by opportunistic pathogens like Micrococcus species on rare occasions. We present a case of Micrococcus sp peritonitis in a 55-year-old female with end-stage kidney disease on continuous cycling peritoneal dialysis for one year who presented with cloudy effluent. Initial treatment against Micrococcus sp with vancomycin, gentamicin, and prophylactic oral nystatin was successful. However, one month later, the patient presented with abdominal pain and dialysate culture again grew Micrococcus sp. Treatment with vancomycin was unsuccessful in resolving culture positivity. The patient was transitioned to hemodialysis for non-medical reasons and then was later restarted on PD without further peritonitis episodes. Micrococcus sp peritonitis in PD poses treatment challenges due to limited guidelines. Intraperitoneal vancomycin is commonly used to target Micrococcus isolates although there is a high incidence of treatment failure. This case report highlights the need for continued reporting to enhance identification, prevention, and patient outcomes in Micrococcus sp peritonitis during PD.


Kidney Failure, Chronic , Peritoneal Dialysis , Peritonitis , Female , Humans , Middle Aged , Vancomycin/therapeutic use , Micrococcus , Peritoneal Dialysis/adverse effects , Peritonitis/drug therapy , Peritonitis/etiology , Kidney Failure, Chronic/complications , Kidney Failure, Chronic/therapy , Anti-Bacterial Agents/therapeutic use
5.
J Antibiot (Tokyo) ; 77(3): 163-169, 2024 03.
Article En | MEDLINE | ID: mdl-38148391

A novel actinobacterial strain, designated as JXJ CY 30 T, was isolated from the phycosphere of Microcystis aeruginosa FACHB-905 (Maf) collected from Lake Dianchi, China. The strain was a Gram-stain-positive, aerobic and coccus-shaped actinobacterium. It had alanine, glutamic acid, aspartic acid, and lysine in the peptidoglycan, and mannose, ribose and arabinose in its cell wall sugars, anteiso-C15:0 and iso-C15:0 as the main cellular fatty acids, MK-7 and MK-8 as the major respiratory quinones, and phosphatidylglycerol, diphosphatidylglycerol, phosphatidylinositol, glycolipid, and an unidentified phospholipid as the polar lipids. The DNA G + C content was 73.08%. Its 16 S rRNA gene sequence shared 99.14%, and 98.75% similarities with Micrococcus flavus DSM 19079 T and M. porci KD337-16T, respectively, and ≤98.41% similarities with other type strains of the genus Micrococcus. It formed independent clade with M. flavus DSM 19079 T on the phylogenetic trees. The digital DNA-DNA hybridization and average nucleotide identity values between strain JXJ CY 30 T and M. flavus DSM 19079 T and M. porci KD337-16T were 48.0% and 92.1%, 25.5% and 83.2%, respectively. These data above indicated that strain JXJ CY 30 T represented a new species of the genus Micrococcus, and the species epithet is proposed as Micrococcus lacusdianchii sp. nov. (type strain JXJ CY 30 T = KCTC 49378 T = CGMCC 1.17508 T). Strain JXJ CY 30 T can potentially provide Maf with various nutrients such as available phosphorus and nitrogen, plant hormones, various vitamins and carotenoids for growth, while it was inhibited by metabolites from its symbiotic algae Maf.


Micrococcus , Phospholipids , Phylogeny , Micrococcus/genetics , Fatty Acids , DNA , RNA, Ribosomal, 16S/genetics , DNA, Bacterial/genetics , Sequence Analysis, DNA , Bacterial Typing Techniques , Vitamin K 2
6.
Arch Microbiol ; 205(11): 351, 2023 Oct 08.
Article En | MEDLINE | ID: mdl-37805972

Lateritic soil is the reddish to brown-colored soil composed mainly of iron or aluminium oxides, hydroxides, or oxyhydroxides. Information on bacteria that inhabit this soil type, their ecological role, and metabolic potential are scarce. We have isolated and partially characterized a bacterial strain BirBP01 from a lead, calcium, and magnesium-rich, oligotrophic subsurface lateritic soil-sample collected from 12-feet deep horizon of a laterite mining pit in Birbhum district, India. The isolate is a biofilm-forming, Gram-positive bacterium having a sarcinae arrangement, mesophilic, slightly alkaliphilic, able to produce amylase, and resistant against multiple heavy-metals. BirBP01 has the ability to bioremediate 51% of Pb, 30% of Zn, and 22% of Cu through biosorption, possibly into the biofilm matrix. The bioremediating ability of the bacterium alleviated the inhibitory effect of heavy-metals on the germination of chickpea (Cicer arietinum L.) seeds. 16S rRNA gene-based phylogenetic analysis revealed that BirBP01 is a member of the genus Micrococcus. It showed more than 99% identity of the 16S rRNA gene sequence, and clustered within the same branch of the phylogenetic tree, with strains of M. yunnanensis, M. endophyticus, and M. luteus. The ability to produce amylase, and bioremediate heavy-metals signify that Micrococcus sp. BirBP01 could be potentially a good candidate for industrial applications, and to clean up heavy-metal contaminated sites.


Metals, Heavy , Soil Pollutants , Micrococcus/genetics , Micrococcus/metabolism , Soil , RNA, Ribosomal, 16S/genetics , Phylogeny , Metals, Heavy/metabolism , Bacteria/genetics , Biofilms , Soil Pollutants/metabolism , Biodegradation, Environmental
7.
Environ Sci Pollut Res Int ; 30(52): 113015-113025, 2023 Nov.
Article En | MEDLINE | ID: mdl-37847368

This study aims to formulate bacterial tablets of cadmium (Cd)-resistant Micrococcus sp. MU1, an indole-3-acetic acid-producer, for soil inoculation to improve Cd phytoremediation by Chlorophytum comosum (Thunb.) Jacques. The viability of Micrococcus sp. MU1 in tablets after storage at room temperature and 4 °C was determined. The ability of Micrococcus sp. tablets and cell suspensions on stimulating growth and Cd accumulation in C. comosum was compared. The results found that the viability of Micrococcus sp. tablets stored at room temperature and 4 °C for 2 months were 29.2 and 97.9%, respectively. After 2 months of growth in pots, the dry biomass weights of C. comosum amended with Micrococcus sp. tablet and cell suspension were greater than that of uninoculated control by 1.4- and 1.3-fold, respectively. Cd concentrations in the roots and shoots of C. comosum inoculated with bacterial tablet and bacterial suspension were not significantly different (p < 0.05) and were greater than that of the uninoculated plants. In addition, plants inoculated with Micrococcus sp. tablet and cell suspension exhibited superior phytoextraction performance, bioaccumulation factor, and translocation factor, indicating equal performance of both bacterial forms on boosting Cd phytoremediation efficiency in C. comosum. These findings suggest that soil inoculation with Micrococcus sp. tablet as a ready-to-use inoculum is a novel approach to promote phytoremediation of C. comosum in Cd-contaminated agricultural soil.


Asparagaceae , Soil Pollutants , Cadmium/analysis , Biodegradation, Environmental , Soil Pollutants/analysis , Plant Roots/chemistry , Soil , Micrococcus
8.
J Ind Microbiol Biotechnol ; 50(1)2023 Feb 17.
Article En | MEDLINE | ID: mdl-37460166

Historically, bacteria of the phylum, Actinobacteria have been a very prominent source of bioactive compounds for drug discovery. Among the actinobacterial genera, Micrococcus has not generally been prioritized in the search for novel drugs. The bacteria in this genus are known to have very small genomes (generally < 3 Mb). Actinobacteria with small genomes seldom contain the well-characterized biosynthetic gene clusters such as those encoding polyketide synthases and nonribosomal peptide synthetases that current genome mining algorithms are optimized to detect. Nevertheless, there are many reports of substantial pharmaceutically relevant bioactivity of Micrococcus extracts. On the other hand, there are remarkably few descriptions of fully characterized and structurally elucidated bioactive compounds from Micrococcus spp. This review provides a comprehensive summary of the bioactivity of Micrococcus spp. that encompasses antibacterial, antifungal, cytotoxic, antioxidant, and anti-inflammatory activities. This review uncovers the considerable biosynthetic potential of this genus and highlights the need for a re-examination of these bioactive strains, with a particular emphasis on marine isolates, because of their potent bioactivity and high potential for encoding unique molecular scaffolds.


Actinobacteria , Micrococcus , Actinobacteria/genetics , Bacteria , Anti-Bacterial Agents/pharmacology , Polyketide Synthases/genetics , Drug Discovery
9.
Sci Rep ; 13(1): 3900, 2023 03 08.
Article En | MEDLINE | ID: mdl-36890143

The present study elucidates identification and characterization of dimethyl phthalate (DMP) degrading novel bacterial strain, Micrococcus sp. KS2, isolated from soil contaminated with municipal wastewater. Statistical designs were exercised to achieve optimum values of process parameters for DMP degradation by Micrococcus sp. KS2. The screening of the ten important parameters was performed by applying Plackett-Burman design, and it delivered three significant factors (pH, temperature, and DMP concentration). Further, response surface methodology involving central composite design (CCD) was implemented to examine mutual interactions among variables and achieve their optimal response. The predicted response indicated that maximum DMP degradation (99.67%) could be attained at pH 7.05, temperature 31.5 °C and DMP 289.19 mg/l. The strain KS2 was capable of degrading up to 1250 mg/l of DMP in batch mode and it was observed that oxygen was limiting factor in the DMP degradation. Kinetic modeling of DMP biodegradation indicated that Haldane model fitted well with the experimental data. During DMP degradation, monomethyl phthalate (MMP) and phthalic acid (PA) were identified as degradation metabolites. This study provides insight into DMP biodegradation process and proposes that Micrococcus sp. KS2 is a potential bacterial candidate to treat effluent containing DMP.


Micrococcus , Phthalic Acids , Micrococcus/metabolism , Phthalic Acids/metabolism , Biodegradation, Environmental , Kinetics , Sewage/microbiology
10.
J Clin Microbiol ; 61(4): e0148422, 2023 04 20.
Article En | MEDLINE | ID: mdl-36946723

Rothia, Kocuria, Arthrobacter, and Pseudoglutamicibacter are bacterial species within the family Micrococcaeae. Knowledge of human infections due to these bacteria is limited. This study aimed to examine features of infections caused by non-Micrococcus Micrococcaeae (NMM). Findings of NMM from blood cultures and other sterile cultures from 2012 to 2021 were identified from the records of the Department of Clinical Microbiology in Region Skåne, Lund, Sweden. Medical records were retrospectively reviewed. True infection was defined as having signs of infection, no other more likely pathogen, and no other focal infection, together with two positive blood cultures or one positive blood culture and an intravascular device. A total of 197 patients with findings of NMM in blood cultures were included. Among adult patients with bacteremia, 29 patients (22%) were considered to have a true infection. Adults with true infection were significantly more likely to have malignancy (69%), leukopenia (62%), and treatment with chemotherapeutics (66%) compared to patients with contaminated samples (24%, 3%, and 8%, respectively) (P < 0.001). A total of 31 patients had findings of NMM in other sterile cultures, and infections were considered true in joints (n = 4), a pacemaker (n = 1), and peritoneal dialysis fluid (n = 1). Infections due to NMM occur but are rare. Growth of NMM in blood cultures should be suspected to be a true infection mainly in immunocompromised patients.


Arthrobacter , Bacteremia , Micrococcaceae , Adult , Humans , Micrococcus , Retrospective Studies , Bacteremia/microbiology
11.
J Appl Microbiol ; 133(5): 3139-3149, 2022 Nov.
Article En | MEDLINE | ID: mdl-35996816

AIM: To reveal the antibacterial mechanism of protocatechuic acid (PCA) against Micrococcus luteus. METHODS AND RESULTS: M. luteus was exposed to PCA, and the antibacterial mechanism was revealed by measuring membrane potential, intracellular ATP and pH levels and transcriptome analysis. PCA induced the membrane potential depolarization of M. luteus, significantly decreased the intracellular ATP and pH levels of M. luteus and disrupted the integrity of the M. luteus cell membrane. Transcriptome analysis showed that PCA induced 782 differentially expressed genes (DEGs) of M. luteus. GO enrichment analysis revealed that the majority of DEGs are involved in pathways of metabolic process, cellular process, biological regulation and transport activity. In addition, PCA inhibited the growth of M. luteus in skimmed milk and extended the shelf life of skimmed milk. CONCLUSION: PCA had good bactericidal activity against M. luteus through the mechanism of cell membrane disruption and metabolic process disorder. SIGNIFICANCE AND IMPACT OF THE STUDY: PCA inhibits the growth of M. luteus in skimmed milk, suggesting that PCA is promising to be used as a novel preservative in food storage.


Gene Expression Profiling , Micrococcus luteus , Anti-Bacterial Agents/pharmacology , Adenosine Triphosphate , Micrococcus
12.
Arch Microbiol ; 204(8): 482, 2022 Jul 14.
Article En | MEDLINE | ID: mdl-35834020

Malathion is widely used as an agricultural insecticide, but its toxic nature makes it a serious environmental contaminant. To screen indigenous bacteria for malathion degradation, a strain MAGK3 capable of utilizing malathion as its sole carbon and energy source was isolated from Pennisetum glaucum agricultural soil. Based on morphological and biochemical characteristics and 16S rDNA sequence analysis, strain MAGK3 was identified as Micrococcus aloeverae. The strain was cultured in the presence of malathion under aerobic and energy-restricting conditions, and it grew well in MSM containing malathion (1000 µl/L), showing the highest specific growth rate at 500 µl/L. Reverse-phase UHPLC-DAD analysis indicated that 100%, 90.48%, 84.27%, 75.46%, 66.65%, and 31.96% of malathion were degraded within 15 days in liquid culture augmented with 50, 100, 200, 300, 500, and 1000 µl/L concentrations of commercial malathion, respectively. Confirmation of malathion degradation to malathion mono, diacids, and phosphorus moiety was performed by Q-TOF-MS analysis, and a pathway of biodegradation was proposed. The influence of co-substrates was also examined to optimize biodegradation further. Kinetic studies based on different models were conducted, and the results demonstrated good conformity with the first-order model. Malathion degradation process by Micrococcus aloeverae was characterized by R2 of 0.95, and the initial concentration was reduced by 50% i.e. (DT50) in 8.11 d at an initial concentration of 500 µl/L. This establishes the Micrococcus sp. as a potent candidate for active bioremediation of malathion in liquid cultures as it can withstand high malathion load and can possibly impact the development strategies of bioremediation for its elimination.


Malathion , Soil Microbiology , Biodegradation, Environmental , Kinetics , Malathion/chemistry , Malathion/metabolism , Malathion/pharmacology , Micrococcus/genetics , Micrococcus/metabolism
13.
J Gen Appl Microbiol ; 68(4): 184-192, 2022 Nov 22.
Article En | MEDLINE | ID: mdl-35598987

Rice (Oryza sativa L.) straw is an agricultural byproduct of high yield, and its disposal by burning has detrimental effect on ecosystem. It has potential as source of fermentable sugars for industrial use; however, it requires effective pretreatment to remove lignin. Bacterial enzymes based pretreatment is advantageous due to their extracellular nature, and tolerance to higher temperature, pH and oxygen limitation. We herein report screening of lignocellulose degradation environment of vermicompost for ligninolytic bacteria, and studying role of Micrococcus unnanensis strain B4 in delignification of rice straw. The bacterium was capable to degrade acid soluble and insoluble lignin; and produced lignin degrading laccase and peroxidase having maximum activity at pH 6.5 and 72 h incubation. Both enzymes exhibited alkaline pH stability, and thermal stability with retention of 100 % activity on pre-incubation at 60 ℃ for 1 h. The enzymes were used for pretreatment of rice straw using chemicals (acetic acid:hydrogen peroxide) pretreatment as reference. Scanning electron microscopy of pretreated rice straw samples showed alteration in morphology with exposure of cellulosic components. Enzymatically pretreated rice straw on saccharification by a commercial cellulase yielded about 400 mg of reducing sugar per gram, comparable to that released on chemical pretreatment. Hence, pretreatment based on M.unnanensis strain B4 and its ligninolytic enzymes can be an alternative to chemical pretreatment for saccharification of rice straw to fermentable sugars.


Cellulase , Oryza , Lignin/metabolism , Oryza/metabolism , Micrococcus/metabolism , Ecosystem , Cellulase/metabolism , Sugars/pharmacology , Hydrolysis
14.
Environ Sci Pollut Res Int ; 29(38): 58201-58211, 2022 Aug.
Article En | MEDLINE | ID: mdl-35359212

Levofloxacin (LFX) is a widely used antibiotic medication. Persistent traces of LFX in water and wastewater may induce bacterial resistance. Photon-driven advanced oxidation processes (AOPs) can assist in attaining complete abatement of LFX for environmental protection. This work benchmarks different solar AOPs based on hydroxyl radical (OH•) and sulphate radical (SO4•-) chemistry. Other oxidant precursors, as radical sources, were used to selectively control the generation of either hydroxyl radical (i.e., H2O2), sulphate radical (i.e., peroxydisulphate (PDS)), or a controlled mixture ratio of both OH•/SO4•- (i.e., peroxymonosulphate (PMS)). The influence of pH on degradation performance was evaluated using unbuffered and buffered solutions. Simulated irradiation/PMS process exhibited a strong pH-dependence attaining partial degradation of ca. 56% at pH 5 up to complete degradation at pH 7. Despite the similitudes on the abatement of target pollutant LFX in pristine solutions, only simulated irradiation/PDS treatment achieved effective abatement of LFX in wastewater samples given the higher selectivity of SO4•-. Toxicity tests were conducted with Escherichia coli (LMG2092) and Micrococcus flavus (DSM1790), demonstrating successful inhibition of the antibiotic character of polluted waters, which would contribute to preventing the development of resistant bacterial strains. Finally, a degradative pathway was suggested from the by-products and intermediates identified by LC-MS. Results demonstrate that the degradation of specific functional groups (i.e., piperazine ring) is associated with the loss of antibacterial character of the molecule.


Wastewater , Water Pollutants, Chemical , Anti-Bacterial Agents/pharmacology , Escherichia coli , Hydrogen Peroxide/chemistry , Hydroxyl Radical/chemistry , Kinetics , Levofloxacin/pharmacology , Micrococcus , Oxidation-Reduction , Sunlight , Wastewater/chemistry , Water Pollutants, Chemical/analysis
15.
Biochim Biophys Acta Proteins Proteom ; 1870(2): 140745, 2022 02 01.
Article En | MEDLINE | ID: mdl-34906734

Bacteriophage endolysins are crucial for progeny release at the end of the lytic cycle. Mycobacteriophage's genomes carry a lysin A essential gene, whose product cleaves the peptidoglycan (PG) layer and a lysin B, coding for an esterase, that cleaves the linkage between the mycolic acids and the arabinogalactan-PG complex. Lysin A mycobacteriophage proteins are highly modular and in gp29 (LysA) of phage TM4 three distinctive domains were identified. By bioinformatics analysis the central module was previously found to be similar to an amidase-2 domain family with an N-acetylmuramoyl -L-alanine amidase activity. We demonstrated experimentally that purified LysA is able to lyse a suspension of Micrococcus lysodeikticus and can promote cell lysis when expressed in E. coli and Mycobacterium smegmatis. After incubation of LysA with MDP (Muramyl dipeptide, N-acetyl-muramyl-L-alanyl-D-isoglutamine) we detected the presence of N-acetylmuramic acid (NAcMur) and L-Ala- D- isoGlutamine (L-Ala-D-isoGln) corroborating the proposed muramidase activity of this enzyme. This protein was stabilized at acidic pH in the presence of Zn consistent with the increase of the enzymatic activity under these conditions. By homology modeling, we predicted that the Zn ion is coordinated by His 226, His 335, and Asp 347 and we also identified the amino acid Glu 290 as the catalytic residue. LysA activity was completely abolished in derived mutants on these key residues, suggesting that the PG hydrolysis solely relies on the central domain of the protein.


Endopeptidases/metabolism , Mycobacteriophages/metabolism , N-Acetylmuramoyl-L-alanine Amidase/metabolism , Peptidoglycan/metabolism , Viral Proteins/metabolism , Computational Biology/methods , Endopeptidases/chemistry , Escherichia coli/metabolism , Galactans , Hydrolysis , Mass Spectrometry/methods , Micrococcus/metabolism , Muramic Acids/metabolism , Mycobacterium smegmatis/metabolism , Viral Proteins/chemistry
16.
Int J Biol Macromol ; 191: 746-752, 2021 Nov 30.
Article En | MEDLINE | ID: mdl-34592219

The carbohydrate recognition domain (CRD) is the key component of C-type lectins (CTLs) with the capacity to recognize and eliminate invading pathogens. Herein, the recombinant proteins of four CRDs identified from the kuruma shrimp, Marsupenaeus japonicus, were produced and purified by an Escherichia coli expression system and affinity chromatography. Bacterial binding and antibacterial assays showed that the four CRDs displayed various bacterial binding and antibacterial activities against different bacteria. Among the four recombinant CRDs, His-CRD2-3 exhibited the broadest spectrum of bacterial binding and antibacterial activities against gram-negative bacteria (Vibrio parahaemolyticus, V. alginolyticus and V. harveyi) and gram-positive bacteria (Staphylococcus aureus and Micrococcus lysodeikticus). Moreover, the four recombinant CRDs showed different capacities to regulate the expression of several immune effector genes (MjCTL3, MjCTL4, MjCTL, Mjily and Mjsty), among which His-CRD2-3 displayed broader and stronger inductive effects on these immune effector genes. This study indicated that the four CRDs participated in immune defense by binding and killing bacteria and regulating the transcription of other immune effector genes. In addition, our results suggested that His-CRD2-3 might be a promising agent for the prevention and treatment of bacteriosis.


Anti-Bacterial Agents/chemistry , Lectins/chemistry , Penaeidae/chemistry , Animals , Anti-Bacterial Agents/pharmacology , Cloning, Molecular , Lectins/genetics , Lectins/pharmacology , Micrococcus/drug effects , Penaeidae/metabolism , Protein Domains , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/pharmacology , Staphylococcus aureus/drug effects , Vibrio/drug effects
17.
Vet Ophthalmol ; 24(4): 326-335, 2021 Jul.
Article En | MEDLINE | ID: mdl-34480390

OBJECTIVE: To describe common bacterial organisms cultured from retrobulbar cellulitis and abscess lesions, in vitro susceptibility patterns, common diagnostic techniques utilized, etiologies encountered, and prevalence of blindness. ANIMALS STUDIED: Thirty-eight dogs diagnosed with retrobulbar cellulitis or abscessation from 2007 to 2017. PROCEDURE: For cases of orbital cellulitis or abscess, signalment, orbital imaging, cytology, histopathology, bacterial culture and susceptibility testing, presence of vision at the initial examination and resolution, and presumed cellulitis/abscess etiology were recorded. RESULTS: Most cases were medically (78.9%) versus surgically managed (18.4%). Most common form of orbital imaging was computed tomography (48.5%) followed by ocular ultrasound (18.2%). Fifteen of eighteen cultures (83.3%) showed growth of aerobic bacterial organisms, anaerobic bacterial organisms, or both. Most common aerobic bacteria were gram-negative bacilli (40.0%) followed by Corynebacterium sp. (26.7%) and α-hemolytic Streptococci sp. (26.7%) but Micrococcus and Bacillus spp. were also identified. Most common anaerobic bacteria were gram-negative bacilli (40.0%). Antibiotics with highest susceptibility patterns included gentamicin, followed equally by amoxicillin/clavulanic acid, cephalothin, chloramphenicol, and imipenem. No bacteria were susceptible to cefovecin. Six cases presented with vision loss due to retrobulbar disease (15.8%). Idiopathic (50%) disease and tooth root abscessation (23.7%) were most commonly diagnosed cause of orbital disease. CONCLUSION: Retrobulbar cellulitis/abscess is a serious and vision-threatening process, which can be effectively managed by broad-spectrum antibiotics such as gentamicin or amoxicillin/clavulanic acid, but not cefovecin. This study identified three organisms that have not been previously reported to be associated with orbital cellulitis (Corynebacterium sp., Bacillus sp. and Micrococcus sp.).


Cellulitis/veterinary , Dog Diseases/diagnosis , Eye Infections, Bacterial/veterinary , Orbital Diseases/veterinary , Animals , Bacillus/isolation & purification , Blindness/microbiology , Blindness/veterinary , Cellulitis/diagnosis , Cellulitis/epidemiology , Cellulitis/therapy , Corynebacterium/isolation & purification , Disease Susceptibility , Dog Diseases/epidemiology , Dog Diseases/microbiology , Dog Diseases/therapy , Dogs , Eye Infections, Bacterial/diagnosis , Eye Infections, Bacterial/epidemiology , Eye Infections, Bacterial/therapy , Female , Male , Micrococcus/isolation & purification , Orbital Diseases/diagnosis , Orbital Diseases/epidemiology , Orbital Diseases/therapy , Southeastern United States/epidemiology , Tomography, X-Ray Computed/veterinary
18.
Inflammation ; 44(6): 2333-2345, 2021 Dec.
Article En | MEDLINE | ID: mdl-34417666

Lipopolysaccharide (LPS) is commonly used in murine sepsis models, which are largely associated with immunosuppression and collapse of the immune system. After adapting the LPS treatment to the needs of locally bred BALB/c mice, the present study explored the protective role of Micrococcus luteus peptidoglycan (PG)-pre-activated vaccine-on-chip technology in endotoxemia. The established protocol consisted of five daily intraperitoneal injections of 0.2 µg/g LPS, allowing longer survival, necessary for a therapeutic treatment application. A novel immunotherapy technology, the so-called vaccine-on-chip, consists of a 3-dimensional laser micro-textured silicon (Si) scaffold loaded with macrophages and activated in vitro with 1 µg/ml PG, which has been previously shown to exert a mild immunostimulatory activity upon subcutaneous implantation. The LPS treatment significantly decreased CD4 + and CD8 + cells, while increasing CD11b + , Gr1 + , CD25 + , Foxp3 + , and class II + cells. These results were accompanied by increased arginase-1 activity in spleen cell lysates and C-reactive protein (CRP), procalcitonin (PCT), IL-6, TNF-a, IL-10, and IL-18 in the serum, while acquiring severe sepsis phenotype as defined by the murine sepsis scoring. The in vivo application of PG pre-activated implant significantly increased the percentage of CD4 + and CD8 + cells, while decreasing the percentage of Gr1 + , CD25 + , CD11b + , Foxp3 + cells, and arginase-1 activity in the spleen of LPS-treated animals, as well as all serum markers tested, allowing survival and rescuing the severity of sepsis phenotype. In conclusion, these results reveal a novel immunotherapy technology based on PG pre-activated micro-texture Si scaffolds in LPS endotoxemia, supporting thus its potential use in the treatment of septic patients.


Immunotherapy/instrumentation , Macrophage Activation/drug effects , Macrophages/drug effects , Peptidoglycan/pharmacology , Sepsis/prevention & control , Spleen/immunology , T-Lymphocyte Subsets/immunology , Tissue Scaffolds , Animals , Cytokines/metabolism , Disease Models, Animal , Equipment Design , Female , Inflammation Mediators/metabolism , Lipopolysaccharides , Macrophages/immunology , Macrophages/metabolism , Male , Mice, Inbred BALB C , Micrococcus/metabolism , Peptidoglycan/isolation & purification , Sepsis/chemically induced , Sepsis/immunology , Sepsis/metabolism , Spleen/metabolism , Surface Properties , T-Lymphocyte Subsets/metabolism
19.
Fish Shellfish Immunol ; 117: 253-261, 2021 Oct.
Article En | MEDLINE | ID: mdl-34418557

The sea urchin Lytechinus variegatus is considered a good candidate for aquaculture, but bacterial diseases are a major challenge in culture conditions. The innate immunological defenses of L. variegatus to bacterial challenges were assessed through hematology parameters, in vitro phagocytosis, lysozyme activity and total plasma protein concentrations in cell-free coelomic fluid. Adult sea urchins were inoculated with Microccocus lysodeikticus, Escherichia coli and Vibrio parahaemolyticus in the cavity coelomic. Filtrated and sterile seawater (FSW) injected and non-injected sea urchins were used as control groups. Righting time, external aspects and behavior of sea urchins were evaluated. Twenty-four hours post-inoculation, we found an increase in the population of colorless spherule cells (CLS), phagocytosis, and humoral responses in sea urchins challenged by bacterial inoculations. Righting time was not affected by the treatments and apparent external signs of disease were not observed at least during 96h post-inoculation. The immunological system of L. variegatus quickly eliminated pathogenic microorganisms. CLS and lysozyme activity cooperate in the immune defenses of L. variegatus, showing an extraordinary efficiency for adjusting the immune defenses under stress caused by microbes. We recommend that the cellular and humoral markers serve as routine tests to monitor health status in sea urchins.


Lytechinus/immunology , Animals , Escherichia coli , Escherichia coli Infections/immunology , Escherichia coli Infections/veterinary , Gram-Positive Bacterial Infections/immunology , Gram-Positive Bacterial Infections/veterinary , Immunity, Innate , Lytechinus/cytology , Lytechinus/microbiology , Micrococcus , Muramidase/immunology , Phagocytosis , Vibrio Infections/immunology , Vibrio Infections/veterinary , Vibrio parahaemolyticus
20.
Fish Shellfish Immunol ; 116: 107-114, 2021 Sep.
Article En | MEDLINE | ID: mdl-34256133

An eight-week investigation was conducted to access the potential impact of dietary watermelon rind powder (WMRP) and L. plantarum CR1T5 (LP) administered individually or in combination on immunity, disease resistance, and growth rate of Nile tilapia fingerlings cultured in a biofloc system. Three hundred twenty fish (average weight 16.57 ± 0.14 g) were distributed into 16 tanks at a rate of 20 fish per tank. The fish were fed different diets: Diet 1 (0 g kg-1 WMRP and 0 CFU g-1 L. plantarum) (control), Diet 2 (40 g kg-1 WMRP), Diet 3 (108 CFU g-1 LP), and Diet 4 (40 g kg-1 WMRP + 108 CFU g-1 LP) for eight weeks. A completely randomized design (CRD) with four replications was applied. Skin mucus, serum immunity, and growth parameters were analyzed every 4 weeks, and a challenge study against S. agalactiae was conducted at the end of the experiment. The findings showed that the inclusion of WMRP + LP, administrated individually or in a mixture, significantly (P<0.05) stimulated growth, skin mucus, and serum immune parameters of Nile tilapia fingerlings compared with the control. The highest values were detected in fish fed the combination of WMRP and LP, as opposed to individual administration of either WMRP or LP, in which no significant differences were detected. Within the challenge study, the relative percent survival (RPS) in Diet 2, Diet 3, and Diet 4 was 48.0%, 52.0%, and 68.0%, respectively. Fish fed 40 g kg-1 WMRP + LP produced significantly higher RPS and protection against S. agalactiae than the other treated groups. Current results suggest that the dual administration of WMRP and LP maybe an effective feed additive for Nile tilapia grown in an indoor biofloc system, capable of improving growth parameters and increasing resistance to S. agalactiae infection.


Citrullus , Lactobacillus plantarum , Plant Preparations/pharmacology , Prebiotics , Synbiotics , Animal Feed , Animals , Aquaculture , Cichlids/blood , Cichlids/growth & development , Cichlids/immunology , Diet/veterinary , Disease Resistance , Leukocyte Count , Micrococcus , Mucus/enzymology , Mucus/immunology , Muramidase/immunology , Peroxidase/immunology , Phagocytosis , Powders , Respiratory Burst , Skin/immunology , Streptococcal Infections/prevention & control , Streptococcus agalactiae
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