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Reactive astrogliosis is a common pathological hallmark of central nervous system (CNS) injury, infection, and neurodegeneration, where reactive astrocytes can be protective or detrimental to normal brain functions. Currently, the mechanisms regulating neuroprotective astrocytes and the extent of neuroprotection are poorly understood. Here, we report that conditional deletion of serum response factor (SRF) in adult astrocytes causes reactive-like hypertrophic astrocytes throughout the mouse brain. TheseSrfGFAP-ERCKO astrocytes do not affect neuron survival, synapse numbers, synaptic plasticity or learning and memory. However, the brains ofSrfknockout mice exhibited neuroprotection against kainic-acid induced excitotoxic cell death. Relevant to human neurodegenerative diseases,SrfGFAP-ERCKO astrocytes abrogate nigral dopaminergic neuron death and reduce β-amyloid plaques in mouse models of Parkinson’s and Alzheimer’s disease, respectively. Taken together, these findings establish SRF as a key molecular switch for the generation of reactive astrocytes with neuroprotective functions that attenuate neuronal injury in the setting of neurodegenerative diseases.

Dopaminergic (DA) neurons in the Olfactory bulb (OB) are involved in odor detection and discrimination. Transcription factor (TF) regulatory network responsible for their fate specification remains poorly understood and the spatial regulation of DA neurons remains elusive. In this study, mice exposed to odor stimulant exhibited specific upregulation of Zinc finger transcription factor of Cerebellum (ZIC) 3 along with Tyrosine Hydroxylase (TH). Stringent co-expression analysis showed ZIC3 and TH dual positive neurons in OB. Genetic manipulation showed ZIC3 to be both essential and sufficient to drive TH expression and essential for odor perception. ZIC3 interacts with ER81 and binds to region encompassing ER81 binding site in DA neurons and is indispensable for TH expression. In midbrain (MB), in the absence of ER81, ZIC3 switches its molecular partner and binds toPitx3promoter-a DA fate determinant. Under ectopic expression of ER81 in MB DA neurons, propensity of ZIC3 binding toPitx3promoter is compromised and its occupancy onThpromoter encompassing ER81 binding site is established, finally culminating in desired TH expression. Together, these findings reveal a unique ZIC3 mediated bimodal regulation of TH in OB and MB to ultimately facilitate DAergic fate.

Heteroresistance is a phenomenon where a subset of bacterial cells shows higher resistance towards antibiotic while the majority are susceptible to the same. Till date, a few reports have been emerged with prevalence of heteroresistance in several bacterial species towards different classes of antibiotics. It is hard to treat heteroresistant bacterial infections as it is difficult in identifying and studying these subpopulations. The organisms with stable heteroresistance can sustain the resistance ability even in the absence of antibiotics while unstable heteroresistance can revert to susceptibility in the absence of antibiotics. The bacterial heteroresistance is due to genetically unsteady amplification of tandem genes or could also be due to physiological mechanism. The major challenge is to detect and distinguish isolates into susceptible or resistant to antibiotics due to instability and low frequency in their appearance that can give misinterpretation in the therapy. This review concentrates on bacterial heteroresistance with their associated mechanisms followed by modified and advanced evaluation procedures. Also, complications allied with the treatment failure due to bacterial heteroresistance and concerns over its prevalence in food chain.

Background & objectives: Pediococcus pentosaceus has been reported to cause clinical infections while it is being promoted as probiotic in food formulations. Antibiotic resistance (AR) genes in this species are a matter of concern for treating clinical infections. The present study was aimed at understanding the phenotypic resistance of P. pentosaceus to macrolide-lincosamide-streptogramin B (MLSB) antibiotics and the transfer of AR to pathogens. Methods: P. pentosacues isolates (n=15) recovered from fermented foods were screened for phenotypic resistance to MLSB antibiotics using disc diffusion and microbroth dilution methods. Localization and transferability of the identified resistance genes, erm(B) and msr(C) were evaluated through Southern hybridization and in vitro conjugation methods. Results: Four different phenotypes; sensitive (S) (n=5), macrolide (M) (n=7), lincosamide (L) (n=2) and constitutive (cMLSB) (n=1) were observed among the 15 P. pentosaceus isolates. High-level resistance (>256 μg/ml) to MLSB was observed with one cMLSB phenotypic isolate IB6-2A. Intermediate resistance (8-16 μg/ml) to macrolides and lincosamides was observed among M and L phenotype isolates, respectively. Cultures with S phenotype were susceptible to all other antibiotics but showed unusual minimum inhibitory concentration (MIC) values of 8-16 μg/ml for azithromycin. Southern hybridization studies revealed that resistance genes localized on the plasmids could be conjugally transferred to Enterococcus faecalis JH2-2. Interpretation & conclusions: The study provides insights into the emerging novel resistance patterns in P. pentosaceus and their ability to disseminate AR. Monitoring their resistance phenotypes before use of MLS antibiotics can help in successful treatment of Pediococcal infections in humans.

Nervous system development and plasticity involve changes in cellular morphology, making morphological analysis a valuable exercise in the study of nervous system development, function and disease. Morphological analysis is a time-consuming exercise requiring meticulous manual tracing of cellular contours and extensions. We have developed a software tool, called SMorph, to rapidly analyze the morphology of cells of the nervous system. SMorph performs completely automated Sholl analysis. It extracts 23 morphometric features based on cell images and Sholl analysis parameters, followed by principal component analysis (PCA). SMorph was tested on neurons, astrocytes and microglia and reveals subtle changes in cell morphology. Using SMorph, we found that chronic 21-day treatment with the antidepressant desipramine results in a significant structural remodeling in hippocampal astrocytes in mice. Given the proposed involvement of astroglial structural changes and atrophy in major depression in humans, our results reveal a novel kind of structural plasticity induced by chronic antidepressant administration.

Nervous system development and plasticity involves changes in cellular morphology, making morphological analysis a valuable exercise in the study of nervous system development, function and disease. Morphological analysis is a time-consuming exercise requiring meticulous manual tracing of cellular contours and extensions. We have developed a software tool, called SMorph, to rapidly analyse the morphology of cells of the nervous system. SMorph performs completely automated Sholl analysis. It extracts 23 morphometric features based on cell images and Sholl analysis parameters, followed by Principal Component Analysis. SMorph is tested on neurons, astrocytes and microglia and reveals subtle changes in cell morphology. Using SMorph, we found that chronic 21-day treatment with antidepressant desipramine results in a significant structural remodeling in hippocampal astrocytes. Given the proposed involvement of astroglial structural changes and atrophy in major depression in humans, our results reveal a novel kind of structural plasticity induced by chronic antidepressant administration.

The objective of this study was to determine the occurrence and distribution of antibiotic resistant (AR) lactic acid bacteria (LAB) in Indian poultry. LAB from poultry farm feces (n = 21) and samples from slaughter houses comprising chicken intestine (n = 46), raw meat (n = 23), and sanitary water (n = 4) were evaluated and compared with those from organic chicken (OC) collected from nearby villages. Screening studies showed 5–7 log units higher erythromycin (ER), tetracycline (TC) and vancomycin (VAN) resistant LAB from conventional poultry chicken (CC) compared to OC. Molecular characterization of isolated cultures (n = 32) with repetitive-PCR profiling and 16S rRNA gene sequencing revealed their taxonomical status as Enterococcus faecium (n = 16), Enterococcus durans (n = 2), Lactobacillus plantarum (n = 10), Lactobacillus pentosus (n = 1) and Lactobacillus salivarius (n = 3). The isolates were found to harbor erm(B), msr(C), msr(A/B), tet(M), tet(L) and tet(K) genes associated with Tn916 and Tn917 family transposons. Expression studies through real-time PCR revealed antibiotic-induced expression of the identified AR genes. In vitro and in vivo conjugational studies revealed transfer of ER and TC resistant (ERR and TCR) genes with transfer frequencies of 10−7 and 10−4 transconjugants recipient−1, respectively. Although no known VAN resistance (VANR) genes were detected, high phenotypic resistance was observed and was transferable to the recipient. From a public health point of view, this study reports Indian poultry as a major source of high levels of AR bacteria contaminating the food chain and the environment. Thus, urgent and determined strategies are needed to control the spread of multiple AR bacteria.

Background & objectives: Pediococcus pentosaceus has been reported to cause clinical infections while it is being promoted as probiotic in food formulations. Antibiotic resistance (AR) genes in this species are a matter of concern for treating clinical infections. The present study was aimed at understanding the phenotypic resistance of P. pentosaceus to macrolide-lincosamide-streptogramin B (MLSB) antibiotics and the transfer of AR to pathogens. Methods: P. pentosacues isolates (n=15) recovered from fermented foods were screened for phenotypic resistance to MLSB antibiotics using disc diffusion and microbroth dilution methods. Localization and transferability of the identified resistance genes, erm(B) and msr(C) were evaluated through Southern hybridization and in vitro conjugation methods. Results: Four different phenotypes; sensitive (S) (n=5), macrolide (M) (n=7), lincosamide (L) (n=2) and constitutive (cMLSB) (n=1) were observed among the 15 P. pentosaceus isolates. High-level resistance (>256 μg/ml) to MLSB was observed with one cMLSB phenotypic isolate IB6-2A. Intermediate resistance (8-16 μg/ml) to macrolides and lincosamides was observed among M and L phenotype isolates, respectively. Cultures with S phenotype were susceptible to all other antibiotics but showed unusual minimum inhibitory concentration (MIC) values of 8-16 μg/ml for azithromycin. Southern hybridization studies revealed that resistance genes localized on the plasmids could be conjugally transferred to Enterococcus faecalis JH2-2. Interpretation & conclusions: The study provides insights into the emerging novel resistance patterns in P. pentosaceus and their ability to disseminate AR. Monitoring their resistance phenotypes before use of MLS antibiotics can help in successful treatment of Pediococcal infections in humans.

Astrocytes play several critical roles in the normal functioning of the mammalian brain including ion homeostasis, synapse formation and synaptic plasticity. Following injury and infection or in the setting of neurodegeneration, astrocytes become hypertrophic and reactive, a process termed astrogliosis. Although acute reactive gliosis is beneficial in limiting further tissue damage, chronic gliosis becomes detrimental for neuronal recovery and regeneration. Several extracellular factors have been identified that generate reactive astrocytes; however, very little is known about the cell-autonomous transcriptional mechanisms that regulate the maintenance of astrocytes in the normal non-reactive state. Herein, we show that conditional deletion of the stimulus-dependent transcription factor, serum response factor (SRF) in astrocytes (SrfGFAP-CKO) results in astrogliosis marked by hypertrophic morphology and increased expression of GFAP, vimentin and nestin. These reactive astrocytes were not restricted to any specific brain region and were seen in both white and grey matter in the entire brain. This astrogliosis persisted throughout adulthood concomitant with microglial activation. Importantly, the Srf mutant mouse brain did not exhibit any cell death or blood brain barrier (BBB) deficits suggesting that apoptosis and leaky BBB are not the causes for the reactive phenotype. The mutant astrocytes expressed more A2 reactive astrocyte marker genes and the SrfGFAPCKO mice exhibited normal neuronal numbers indicating that SRF-deficient gliosis astrocytes are not neurotoxic. Together our findings suggest that SRF plays a critical role in astrocytes to maintain them in a non-reactive state. SIGNIFICANCE STATEMENT In response to CNS injury, infection and in neurodegeneration, astrocytes undergo complex structural and physiological changes termed as reactive gliosis. Currently, the molecular mechanisms that regulate the non-reactive state of the astrocytes are poorly understood. We report that the SRF transcription factor is required for the maintenance of astrocytes in the non-reactive state such that its conditional deletion in astrocytes results in widespread reactive astrogliosis. The SRF-deficient reactive astrocytes are persistent, non-proliferating and are not caused by cell death or impaired blood brain barrier integrity. In this regard, SRF regulates reactive astrocyte generation in the mammalian brain in a cell-autonomous manner.

Background Despite the growing importance of probiotics apparent health benefits, an impediment to the use of new probiotic cultures is their safety. Hence there is a need to strictly examine the biosafety as well as health benefits of probiotics in in vivo model systems. Results In this study, two lactic acid bacterial (LAB) cultures Lactobacillus fermentum NCMR 2826 and FIX proven for their in vitro probiotic properties were investigated for their in vivo safety in Wistar rats. An acute toxicity study (14 days) with a high dose of biomass (1016 colony-forming units (CFU) mL-1 ) followed by a subchronic test for 13 weeks with oral feeding of the probiotic cultures in three different doses (107 , 108 and 1010 CFU mL-1 ) on a daily basis revealed the safety of the L. fermentum cultures. The probiotic feeding had no toxic effects on survival, body weight and food consumption with any of the dosages used throughout the treatment period. No statistically significant changes in relative organ weights and serum biochemical and hematological indices were found between the control and the probiotic fed animals. In addition to the safety attributes, the L. fermentum culture fed rats showed reduced serum cholesterol levels, macrovesicular steatosis and hepatocyte ballooning compared with control animals. Further, quantification of intestinal microbiota using real-time polymerase chain reaction (PCR) analysis from animal feces indicated a significant increase and stability of Lactobacillus and Bifidobacterium counts but a decrease of Escherichia coli numbers. Conclusion This study of safety and beneficial features highlights the use of the two native L. fermentum isolates as potential probiotic food supplements. © 2019 Society of Chemical Industry.

Three strains of Lactobacillus comprising Lactobacillus salivarius (CHS-1E and CH7-1E) and Lactobacillus reuteri (CH2-2) previously isolated from chicken meat were analyzed for their transferability of antibiotic resistance (AR) genes to pathogenic strains under in vivo, in vitro, and during food fermentation. For in vivo model, Albino Wistar rats were inoculated with 1010 CFU/g/ml of Enterococcus faecalis JH2-2 (recipient). After 7 days, either of two donors L. salivarius CH7-1E or L. reuteri [harbouring erythromycin and tetracycline resistance genes] were introduced at a concentration of 109 CFU/ml daily for 1 week. Two days after donor introduction, there was a stable increase in the number of transconjugants in the animal faeces from 102 to 103 CFU/g and presented erm(B), tet(M), tet(L) and tet(W) in their genome like donor strains. Similar observations were made with in vitro filter mating between CHS-1E, CH2-2 and CH7-1E and E. faecalis JH2-2 with transfer frequencies of 1 × 10−4, 3.8 × 10−3 and 2 × 10−3 per donor cell respectively. With the results obtained in vivo and in vitro, the AR transferability of donor strains was estimated during food fermentation (chicken sausage, fermented milk or idli batter) with pathogenic recipient strains added as contaminants. At the end of mating period, phenotypic resistance to erythromycin and tetracycline in Listeria monocytogenes and Yersinia enterocolitica strains was observed. This study showed the ability of food borne Lactobacillus in diffusing their AR traits in diverse natural environments increasing their concern of AR dissemination in the food chain when used as food additives and/or probiotics.

Antibiotic resistance is a growing problem in clinical settings as well as in food industry. Lactic acid bacteria (LAB) commercially used as starter cultures and probiotic supplements are considered as reservoirs of several antibiotic resistance genes. Macrolide-lincosamide-streptogramin (MLS) antibiotics have a proven record of excellence in clinical settings. However, the intensive use of tylosin, lincomysin and virginamycin antibiotics of this group as growth promoters in animal husbandry and poultry has resulted in development of resistance in LAB of animal origin. Among the three different mechanisms of MLS resistance, the most commonly observed in LAB are the methylase and efflux mediated resistance. This review summarizes the updated information on MLS resistance genes detected and how resistance to these antibiotics poses a threat when present in food grade LAB.

Lactic acid bacteria (LAB) resistant to erythromycin were isolated from different food samples on selective media. The isolates were identified as Enterococcus durans, Enterococcus faecium, Enterococcus lactis, Enterococcus casseliflavus, Lactobacillus salivarius, Lactobacillus reuteri, Lactobacillus plantarum, Lactobacillus fermentum, Pediococcus pentosaceus and Leuconostoc mesenteroides. Of the total 60 isolates, 88 % harbored the ermB gene. The efflux gene msrA was identified in E. faecium, E. durans, E. lactis, E. casseliflavus, P. pentosaceus and L. fermentum. Further analysis of the msrA gene by sequencing suggested its homology to msrC. Resistance to tetracycline due to the genes tetM, tetW, tetO, tetK and tetL, alone or in combination, were identified in Lactobacillus species. The tetracycline efflux genes tetK and tetL occurred in P. pentosaceus and Enterococcus species. Since it appeared that LAB had acquired these genes, fermented foods may be a source of antibiotic resistance.