AggLb Is the Largest Cell-Aggregation Factor from Lactobacillus paracasei Subsp paracasei BGNJ1-64, Functions in Collagen Adhesion, and Pathogen Exclusion In Vitro

Eleven Lactobacillus strains with strong aggregation abilities were selected from a laboratory collection. In two of the strains, genes associated with aggregation capability were plasmid located and found to strongly correlate with collagen binding. The gene encoding the auto-aggregation-promoting protein (AggLb) of Lactobacillus paracasei subsp. paracasei BGNJ1-64 was cloned using a novel, wide-range-host shuttle cloning vector, pAZILSJ. The clone pALb35, containing a 11377-bp DNA fragment, was selected from the SacI plasmid library for its ability to provide carriers with the aggregation phenotype. The complete fragment was sequenced and four potential ORFs were detected, including the aggLb gene and three surrounding transposase genes. AggLb is the largest known cell-surface protein in lactobacilli, consisting of 2998 aa (318,611 Da). AggLb belongs to the collagen-binding superfamily and its C-terminal region contains 20 successive repeats that are identical even at the nucleotide level. Deletion of aggLb causes a loss of the capacity to form cell aggregates, whereas overexpression increases cellular aggregation, hydrophobicity and collagen-binding potential. PCR screening performed with three sets of primers based on the aggLb gene of BGNJ1-64 enabled detection of the same type of aggLb gene in five of eleven selected aggregation-positive Lactobacillus strains. Heterologous expression of aggLb confirmed the crucial role of the AggLb protein in cell aggregation and specific collagen binding, indicating that AggLb has a useful probiotic function in effective colonization of host tissue and prevention of pathogen colonization

Preliminary investigation of polycyclic aromatic hydrocarbons in street dust of Pančevo, Serbia

Polycyclic aromatic hydrocarbons (PAHs) is a common name for a large group of organic compounds with two or more fused aromatic rings. PAHs are well known for their toxicity [1,2]. Because of that they keep attracting great interest on many environmental studies. PAHs can have natural or anthropogenic origin. In nature, PAHs are produced during fires and volcanic eruptions, during formation of petroleum and coal, as a result of diagenesis of organic matter and by biosynthesis. The main anthropogenic sources of PAHs are processing of coal and crude oil, combustion and vehicle traffic [3]. Urban street dust is the sink of PAHs from different sources [4,5]. There is a general agreement that the higher amounts of PAHs in street dusts, the higher human exposure risk to these pollutants via inhalation, ingestion, or dermal contact [6]. The aim of this study was a preliminary investigation of PAHs in street dust of the Pančevo city, Serbia. A representative composite street dust sample was collected from different locations in the Pančevo city. Using a set of stainless steel sieves the particles were fractioned into three sizes with diameters of: < 63 μm, 63 – 250 μm, and 250 – 500 μm. The street dust particles were extracted and the extracts were cleaned up and fractionated using column chromatography. PAHs were analysed by gas chromatography – mass spectrometry (GC-MS) in a selected ion monitoring (SIM) mode. The ions monitored were: m/z = 128 (naphthalene), m/z = 152 (acenaphthylene), m/z = 154 (acenaphthene), m/z = 166 (fluorene), m/z = 178 (phenanthrene and anthracene), m/z = 202 (fluoranthene and pyrene), m/z = 228 (benzo[a]anthracene and chrysene), m/z = 252 (benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[j]fluoranthene, benzo[e]pyrene and benzo[a]pyrene), m/z = 276 (indeno[1,2,3-c,d] pyrene and benzo[g,h,i]perylene), and m/z = 278 (dibenzo[a,h]anthracene). The results showed that all PAHs investigated were present in the analysed composite sample. Furthermore, a non-target screening of the extracts indicated presence of some other environmentally important aromatic hydrocarbons such as: acetophenone, benzophenone, biphenyl and retene. The analysis of the distribution pattern of the PAHs different in size in different size fractions revealed a possible size fractionation of three- and tetracyclic PAHs. These results indicate that PAHs represent an important component in the street dust of the Pančevo city and point to the need for future investigation of the dust samples from this locality

Matematičko modelovanje fenomena transporta Cs+ iz solidifikovane istrošene jonoizmenjivačke smole

The aim of the study was to assess Cs+ ions transport phenomena from solidified spent ion exchange resin by mathematical modeling. The experimental results comparison was obtained by Hespe's Standard Leaching Method. For the leaching prediction rate as a function of time, diffusion and semi-empirical models were used. Due to the presence of spent ion exchange resin, the cement matrix absorbed a larger amount of water, swelled, and degraded. This phenomenon caused a significantly lower value of mechanical resistance to pressure. Also, through the increase of bentonite and zeolite content, the cement matrix decreased its mechanical resistance. The retention of cesium ions in the cement matrix was low and they were leached during the early phase of the investigation. The diffusion coefficient, De, decreased by three orders of magnitude with the addition of zeolite and bentonite in the cement matrix. Linear regression of experimental Cs+ leaching results, under static conditions, displayed that the semi-empirical parameter K3 than the absolute values of the parameters K2 and K1. Therefore, the contribution of matrix dissolution to the total radionuclides transport was irrelevant to the prevailing share of diffusion and surface washing processes

The presence of prtP proteinase gene in natural isolate Lactobacillus plantarum BGSJ3-18

Aims: The study of proteolytic activity and examination of proteinase gene region organization in proteolytically active Lactobacillus plantarum strains from different natural sources. Methods and Results: A set of 37 lactobacilli was distinguished by using multiplex PCR assay. Results showed that 34 strains were Lact. plantarum and three of them were Lact. paraplantarum. The examination of proteolytic activity revealed that 28 Lact. plantarum and two Lact. paraplantarum hydrolyse beta-casein. Further analyses of all proteolytically active Lact. plantarum with primers specific for different types of CEPs demonstrated that strain BGSJ3-18 has prtP catalytic domain as well as prtP-prtM intergenic region showing more than 95% sequence identity with the same regions present in Lact. paracasei, Lact. casei and L. lactis. No presence of prtB, prtH or prtR proteinase genes was detected in any of tested Lact. plantarum strains. Conclusions: One out of 28 analysed Lact. plantarum strains harbours the prtP-like gene. The other proteolytically active Lact. plantarum probably possesses a different type of extracellular proteinase(s). Significance and Impact of the Study: It is the first report about the presence of the prtP-like gene in Lact. plantarum, which illustrates the mobility of this gene and its presence in different species

Comparative analysis of antimicrobial and proteolytic activity of lactic acid bacteria isolated from Zlatar cheese

Tradicionalni zlatarski sir pripada grupi belih, polutvrdih sireva proizvedenih u domaćinstvu. Sir se proizvodi od nepasterizovanog kravljeg mleka bez dodavanja bilo kakvih poznatih starter kultura. Ukupno je izolovano 253 Gram pozitivnih i katalaza negativnih bakterija mlečne kiseline (BMK). Rezultati su pokazali da 70 od 253 analiziranih izolata proizvodi antimikrobna jedinjenja poznatih kao bakteriocini. Većina izolata koji pripadaju rodovima Lactococcus i Enterococcus, kao i izolati vrsta Lactobacillus plantarum i Lb. brevis ne sintetišu ekstracelularne proteinaze. Nasuprot njima, izolati prodvrste Lb. paracasei subsp. paracasei pokazuju veoma dobru proteolitičku aktivnoist. Pokazano je da ne postoji korelacija između dobre proteolitičke i antimikrobne aktivnosti u većini izolata.Traditional artisan Zlatar cheese belongs to the group of white, semi hard home-made cheeses, which are produced from no pasteurized cow's milk, without addition of any known bacterial starter culture. In total, 253 Gram-positive and catalase negative lactic acid bacteria (LAB) were isolated. Results showed that 70 out of 253 analyzed isolates produced antimicrobial compounds known as bacteriocins. Most isolates from genera Lactococcus and Enterococcus, and isolates belonging to species Lactobacillus plantarum and Lb. brevis, do not synthesize extracellular proteinase. In contrast, isolates from subspecies Lb. paracasei subsp. paracasei showed very good proteolytic activity. It was observed that good proteolytic activity of isolates was not in correlation with their good antimicrobial activity in the most of isolates

Lactobacilli hydrolysis of cows' milk proteins abrogates their humoral immunoreactivity in patients with immune-mediated diseases

The level of humoral immunoreactivity to total cows' milk proteins (TCMP) in sera of patients suffering from recurrent oral ulcerations, gastrointestinal diseases or haematological malignancies was investigated. TCMP were also hydrolysed with two different species of lactobacilli and dramatic changes in the levels of specific IgG and IgE were found with statistically significant decreases in the levels of specific antibodies in sera from all patient groups. The levels below cut-off values of IgG specific for TCMP hydrolysates were detected in sera from all patients, while values of IgE for hydrolysates obtained with Lactobacillus helveticus BGRA43 and Lactobacillus zeae LMG17315were below cut-off in 85% and 97% of patients, respectively. Competitive ELISA confirmed the specificity of antibodies for immunogenic TCMP epitopes, demonstrating that lactobacilli hydrolyse TCMP by degrading immunogenic epitopes, and could therefore be used in processing of milk proteins to obtain products suitable for patients with altered immune response on TCMP.This is the peer reviewed version of the paper: Vukotic, G., Matic, I., Begovic, J., Besu, I., Kojic, M., Djokic, J., Juranic, Z., & Strahinic, I. (2016). Lactobacilli hydrolysis of cows’ milk proteins abrogates their humoral immunoreactivity in patients with immune-mediated diseases. International Dairy Journal, 63, 1–7. [https://doi.org/10.1016/j.idairyj.2016.07.009

Retraction notice to: Kojić, M.; Lozo, J.; Jovčić, B.; Strahinić, I.; Fira, Đ.; Topisirović, L. A Successful Use of a New Shuttle Cloning Vector pA13 for the Cloning of the Bacteriocins BacSJ and Acidocin 8912. Archives of Biological Sciences 2010, 62 (2), 231–243. https://doi.org/10.2298/ABS1002231K.

Retraction notice to: Kojić, M.; Lozo, J.; Jovčić, B.; Strahinić, I.; Fira, Đ.; Topisirović, L. A Successful Use of a New Shuttle Cloning Vector pA13 for the Cloning of the Bacteriocins BacSJ and Acidocin 8912. Archives of Biological Sciences 2010, 62 (2), 231–243. [https://doi.org/10.2298/ABS1002231K]

Characterization and antimicrobial activity of natural isolate Lactococcus lactis subsp. lactis BGSM1-19

Soj Lactococcus lactis subsp. lactis BGSM1-19, izolovan iz sira tradicionalno proizvedenog u domaćinstvu, sintetiše dva bakteriocina: bakteriocin BacSMa koji pripada grupi laktokokcina B i bakteriocin BacSMb koji pokazuje visoku homologiju sa lakticinom RM. Čišćenjem plazmida, sa relativno niskim prinosom (0,33%), dobijene su dve grupe derivata: BacSMa- BacSMas derivat i BacSMa- BacSMas, BacSMb-, BacSMbs.. Sinteza bakteriocina je ispitivana tokom logaritamske faze rasta pri čemu je utvrđen maksimum proizvodnje u kulturi staroj 8 sati gajenoj na temperaturi od 30oC, što odgovara ranoj stacionarnoj fazi rasta. Biohemijska karakterizacija je ukazala da soj BGSM1-19 zadržava antimikrobnu aktivnost u opsegu pH vrednosti od 1 do 12 kao i posle tretmana na 100 oC u trajanju od 15 minuta. Utvrđeno je da se antimikrobna aktivnost potpuno gubi nakon tretmana različitim proteolitičkim enzimima. Soj BGSM1-19 poseduje pet plazmida. U eksperimentima čišćenja od plazmida utvrđeno je da se geni za sintezu i imunost na bakteriocine nalaze na plazmidima. Pored toga, BGSM1-19 pokazuje antimikrobnu aktivnost na ispitivane patogene bakterije kao što su Salmonella paratyphi, Micrococcus flavus, Pseudomonas aeruginosa i Staphylococcus aureus.The strain Lactococcus lactis subsp. lactis BGSM1-19, isolated from traditionally homemade white cheese, produces two bacteriocins: lactococcin B-like bacteriocin named bacteriocin BacSMa and bacteriocin BacSMb which have shown similarity with lacticin RM. Plasmid curing resulted in a low yield (0.33%) of BacSMa- BacSMas and BacSMa- BacSMas, BacSMb-, BacSMbs derivatives. The bacteriocin biosynthesis was observed in the logarithmic phase of growth and the production plateau was reached after 8 h of incubation at 30oC, when the culture entered the early stationary phase. Biochemical characterization showed that strain BGSM1-19 retained antimicrobial activity within the pH range of 1 to 12 or after treatment at 100oC for 15 min. However, bacteriocin activity was completely lost after treatment with different proteolytic enzymes. The strain BGSM1-19 contains five plasmids. Plasmid curing indicated that genes coding for bacteriocins synthesis and immunity seem to be located on plasmids. BGSM1-19 exhibited antimicrobial activity against some pathogenic bacteria such as Salmonella paratyphi, Micrococcus flavus, Pseudomonas aeruginosa and Staphylococcus aureus