Activity patterns of central amygdala neurons in a mouse model of narcolepsy

Narcolepsy is a disorder of unstable wake and sleep states caused by the lack of orexin neurons which degenerate most likely as a consequence of an autoimmune process. The state instability of narcolepsy includes rapid eye movement (REM) sleep intruding into wake in the form of dream-like hallucinations and cataplexy, muscle paralysis (atonia) much like occurs in REM sleep. In mice lacking orexin peptides, cataplexy is also observed with similar presentation as in humans of muscle paralysis during wakefulness which is often triggered by positive emotions. Prior research showed that the activation of the central amygdala is sufficient to promote cataplexy in a mouse model of narcolepsy. The central amygdala (CeA) contains a variety of neuronal types, and we hypothesize that γ-aminobutyric acid (GABA)-ergic neurons expressing the oxytocin receptor (OTR) mediate cataplexy as these neurons project to a known REM sleep atonia-regulating region, the ventrolateral periaqueductal gray (vlPAG)/lateral pontine tegmentum (LPT), and, as oxytocin (OT) sensitive neurons in the amygdala, likely participate in emotional processing and social behavior. In this study, we used fiber photometry to investigate the behavior of these neurons in response to social and rewarding stimuli, during emotion-triggered cataplexy, and across arousal states in an effort to define their potential role in emotion-triggered cataplexy. Initial recordings were conducted at too low an excitation light power to stimulate the green fluorescent calcium indicator, GCaMP6s, but were useful in optimizing MATLAB analysis and behavioral tests later done at higher LED power. The second series of recordings with higher excitation light power and better signal to noise ratio, showed increased activity in response to social interaction and reward, prior to REM transitions, and decreased activity during cataplexy confirming patterns seen in initial recordings. In recordings with higher excitation light, these responses appear to occur before interaction with stimulus mice or reward stimulus. In the future, additional recordings with a higher signal to noise ratio will be needed to confirm these results. In conclusion, responses of CeA-OTR neurons to social and rewarding stimuli, cataplexy, and at REM transitions are in support of a possible role of these neurons in emotion-triggered cataplexy which can be tested using additional methods, such as optogenetics

Cloning and expression of a novel lactococcal aggregation factor from Lactococcus lactis subsp. lactis BGKP1

<p>Abstract</p> <p>Background</p> <p>Aggregation may play a main role in the adhesion of bacteria to the gastrointestinal epithelium and their colonization ability, as well as in probiotic effects through co-aggregation with intestinal pathogens and their subsequent removal. The aggregation phenomenon in lactococci is directly associated with the sex factor and lactose plasmid co-integration event or duplication of the cell wall spanning (CWS) domain of PrtP proteinase.</p> <p>Results</p> <p><it>Lactococcus lactis </it>subsp. <it>lactis </it>BGKP1 was isolated from artisanal semi-hard homemade cheese and selected due to its strong auto-aggregation phenotype. Subsequently, non-aggregating derivative (Agg^-) of BGKP1, designated as BGKP1-20, was isolated, too. Comparative analysis of cell surface proteins of BGKP1 and derivative BGKP1-20 revealed a protein of approximately 200 kDa only in the parental strain BGKP1. The gene involved in aggregation (<it>aggL</it>) was mapped on plasmid pKP1 (16.2 kb), cloned and expressed in homologous and heterologous lactococci and enterococci. This novel lactococcal aggregation protein was shown to be sufficient for cell aggregation in all tested hosts. In addition to the <it>aggL </it>gene, six more ORFs involved in replication (<it>repB </it>and <it>repX</it>), restriction and modification (<it>hsdS</it>), transposition (<it>tnp</it>) and possible interaction with mucin (<it>mbpL</it>) were also located on plasmid pKP1.</p> <p>Conclusion</p> <p>AggL is a new protein belonging to the collagen-binding superfamily of proteins and is sufficient for cell aggregation in lactococci.</p

Post-translational regulation of the RpoS and PsrA genes in pseudomonas putida WCS358: The role of ClpXP protease

RpoS i PsrA proteini su ključni transkripcioni regulatori koji se kod pseudomonada aktiviraju u odgovoru na stacionarnu fazu rasta. Cilj ove studije bio je utvrđivanje uloge ClpXP (ATP zavisna serin proteaza) u stabilnosti RpoS i PsrA proteina tokom različitih faza rasta u Pseudomonas putida WCS358. "Western blot" analiza proteinskih ekstrakata P. putida WCS358 i P. putida WCS358 clpX::Km iz rane eksponencijalne, Kasne eksponencijalne i stacionarne faze rasta, sa antitelima na RpoS i PsrA, pokazala je da ClpXP degraduje RpoS i PsrA u ranoj eksponencijalnoj fazi rasta.The RpoS and PsrA proteins are key transcriptional regulators that are activated in response to the stationary phase of growth in pseudomonads. This study was designed to establish whether ClpXP (ATP-dependent serine protease) regulates levels of RpoS and PsrA in Pseudomonas putida WCS358. Western blot analysis of P. putida WCS358 protein extracts from the early exponentianl, late exponential, and stationary phases of growth with antibodies against RpoS and PsrA revealed that these proteins are degraded by ClpXP in the early exponential phase of growth. The obtained results demonstrate a role for ClpXP protease in post-translational regulation of proteins encoded by the rpoS and psrA genes in Pseudomonas spp

Dynamics of sodium dodecyl sulfate utilization andantibiotic susceptibility of strain Pseudomonas sp. ATCC19151

Poznato je da soj Pseudomonas sp. ATCC19151 poseduje gen koji kodira Potencijalnu alkilsulfatazu. U ovom radu analizirana je sposobnost rasta ovog soja u minimalnom medijum usa različitim koncentracijama natrijum dodecilsulfata (0.5, 0.75 i 1 %) kao jedinim izvorom ugljenika. Pokazano je da Pseudomonas sp. ATCC19151 ispoljava najbolji rast uminimalnom medijumusa 0.5 % natrijum dodecil sulfata, te je stoga ova koncentracija uzeta kao optimalna za testiranje dinamike korišćenja natrijum dodecil sulfata tokom različitih faza rasta. Dinamika korišćenja natrijum dodecil sulfata podudarala se sa rastom kulture. Pored toga u cilju detaljnije karakterizacije soja, analizirana je i osetljivost Pseudomonas sp. ATCC19151 na antibiotike. Pokazano je da je analizirani soj rezistentan na šest (ampicilin, tetraciklin, hloramfenikol, tobramicin, nalidiksičnukiselinui gentamicin) od devet analiziranih antibiotika.Pseudomonas sp. ATCC19151 harbors a gene encoding a putative alkylsulfatase (sdsA). Here we report a growth ability of this strain in minimal media containing 0.5, 0.75, and 1% sodium dodecyl sulfate as the sole carbon source. The most prominent growth was detected for the minimal medium with 0.5% SDS, so this concentration of SDS was used to monitor Pseudomonas sp. ATCC19151 SDS biodegradation dynamics. Bacterial growth coincided with the disappearance of SDS. Antibiotic susceptibility was tested as well. Pseudomonas sp. ATCC19151 was resistant to six out of nine tested antibiotics, including ampicillin, tetracycline, chloramphenicol, tobramycin, nalidixic acid, and gentamycin

The clinical isolate Pseudomonas aeruginosa MMA83 carries two copies of the blaNDM-1 gene in a novel genetic context.

The genetic context of the bla(NDM-1) gene in the genome of Pseudomonas aeruginosa MMA83 was investigated. Sequencing of the cosmid selected for the bla(NDM-1) gene revealed the presence of two bla(NDM-1) copies in the genome of P. aeruginosa MMA83 in a unique genetic environment. Additionally, mating assays, DNA-DNA hybridization, and an S1 nuclease assay strongly suggest that the bla(NDM-1) gene in P. aeruginosa MMA83 is chromosome borne

Analysis of natural isolates of Lactobacilli resistant to bacteriocin nisin

Kolekcija bakterija mlečne kiseline (BMK) je napravljena od mikroorganizama izolovanih iz fermentisanih mlečno-kiselinskih proizvoda dobijenih na tradicionalan način. Iz kolekcije 51 izolat je identifikovan kao Lactobacillus sp. Svi izolovani laktobacili pripadaju grupi mezofilnih sojeva koji dobro rastu na temperaturama od 15°C i 30°C, a ne rastu na temperaturi od 45°C. Testiranje sposobnosti rasta u prisustvu nizina pokazalo je da su izolati BGCGK4, BGHN40, BGBUK2-8, BGBUK2-7 i BGBUK2-16 rezistentni na bakteriocin nizin. U eksperimentu određivanja minimalne inhibitorne koncentracije (MIK) za nizin pokazano je da je najrezistentniji izolat Lactobacillus sp. BGCGK4. Izolat BGBUK2-16, determinisan kao Lactobacillus paracasei subsp. paracasei, produkuje bakteriocin označen kao Bac217 i pokazuje rezistenciju na 8000 IU/ml. Čišćenjem plazmida iz soja BGBUK2-16 dobijena su 2 derivata označena kao BGBUK2-16/K2 i BGBUK2-16/K4. Derivat BGBUK2-16/K2 zadržao je rezistenciju na Bac217 i nizin, ali je izgubio sposobnost sinteze Bac217, dok je derivat BGBUK2-16/K4 pored gubitka sposobnosti sinteze Bac217 postao senzitivan na Bac217 i nizin. Prirodno rezistentni laktobacili se mogu iskoristiti za pripremanje starter kultura u kombinaciji sa nizinom kao konzervansom u cilju kontrolisane mlečno-kiselinske fermentacije.The collection of lactic acid bacteria (LAB) was made by isolation of microorganisms from fermented products traditionally manufactured in different geographical regions (high mountains, river valleys, seaside, etc). Among collected LAB, 51 isolates were identified as Lactobacillus sp. Results showed that all isolated lactobacilli were mesophilic strains, since they grew at 15°C and 30°C but not at 45°C. Testing the ability of isolated lactobacilli to grow in the presence of nisin revealed that Lactobacillus sp. isolates designed BGCGK4, BGHN40, BGBUK2-8, BGBUK2-7 and BGBUK2-16 were resistant to nisin. Determination of the minimal inhibitory concentrations (MIC) for nisin revealed that the most resistant isolate was Lactobacillus sp. BGCGK4. Isolate BGBUK2-16, determined as Lactobacillus paracasei subsp. paracasei, produces bacteriocin, named Bac217 and showed a resistance to 8000 IU/ml of nisin. Plasmid curing of BGBUK2-16 resulted in derivatives BGBUK2-16/K2 and BGBUK2-16/K4. Derivative BGBUK2-16/K2 retained resistance to Bac217 and nisin, but lost the ability to synthesise Bac217. Derivative BGBUK2-16/K4 lost concomitantly the resistance to both Bac217 and nisin

Characterization and antimicrobial activity of vaginal lactobacillus isolate

The aim of this study was to investigate the probiotic potential of bacteriocin-producing lactobacilli strain Lactobacillus plantarum G2 isolated from the vaginal mucus of healthy women. The antimicrobial effect of G2 was confirmed in the mixed culture with pathogenic Escherichia coli, Staphylococcus aureus, Salmonella abony and Pseudomonas aeruginosa, while bacteriocine activity was detected against S. aureus and S. abony only. The strain showed an excellent survival rate in low pH and in the presence of bile salts. The percentage of adhered cells of L. plantarum G2 to hexadecane was 63.85±2.0 indicating the intermediate hydrophobicity

Human vaginal Lactobacillus rhamnosus harbor mutation in 23S rRNA associated with erythromycin resistance

Little is known about the diversity and distribution of resistance determinants in human commensal bacteria. The aim of this study was to determine the molecular mechanism responsible for high-level erythromycin resistance among five human vaginal Lactobacillus rhamnosus isolates. PCR screening for the presence of ermA, ermB and ermC methylase genes revealed no determinants responsible for detected erythromycin resistance. Therefore, sequences of 23S rRNA genes from L. rhamnosus strains were studied by PCR-RFLP analysis and sequencing of 23S rRNA genes. According to the results, in all erythromycin-resistant L. rhamnosus strains, the presence of a A - gt G transition mutation at position 2058 was discovered. Additionally, the isolates exhibited heterozygosity for the A2058/G2058 mutation among 23S rRNA gene copies. Presumably, the greatest number of mutated 23S rRNA operons was observed for the L. rhamnosus BGHV1' strain that also had the highest MIC for erythromycin (MIC = 2048 mu g mL(-1)). This study reports the presence of transition mutations in the V region of 23S rRNA genes that most probably account for high-level erythromycin resistance observed for the first time in human vaginal lactobacilli.Peer-reviewed manuscript: [https://imagine.imgge.bg.ac.rs/handle/123456789/1618

Probiotic features of two oral Lactobacillus isolates

In this study, we checked lactobacilli strains of human origin for their potential as probiotic. Samples were collected from oral mucosa of 16 healthy individuals, out of which twenty isolates were obtained and two of them were selected and identified as Lactobacillus plantarum (G1) and L. casei (G3). Both isolates exhibited antagonistic action towards pathogenic microorganisms such as Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Salmonella abony, and Clostridium sporogenes, but not on the growth of Candida albicans. The bacteriocin activity against Staphylococcus aureus ATCC 6358-P was shown only by L. plantarum G1. Moreover, the isolates G1 and G3 showed good viability in the acid gastric environment and in the gut environment containing bovine bile salts. The viability of G1 and G3 isolates in the gastrointestinal tract, and the adhesion to the intestinal mucosa were also confirmed in vivo. The biochemical tests of blood samples revealed lower levels of serum triglycerides and cholesterol, as well as reduced activity of alkaline phosphatase in all lactobacilli-treated Wistar rats, compared to control ones. No toxicity for NMRI Ham mice was observed. According to our experimental results, these findings imply that L. plantarum G1 and L. casei G3 could be characterized as potential probiotics