Papers by Dr. Ashutosh Sharma
Agronomy
Superfruit guava (Psidium guajava L.) is one of the healthiest fruits due to its high antioxidant... more Superfruit guava (Psidium guajava L.) is one of the healthiest fruits due to its high antioxidant dietary fiber and vitamin content. However, the growth and development of this plant are severely affected by salinity stress, mostly at the seedling stage. MicroRNAs (miRNAs) are small, noncoding, endogenous, highly conserved RNA molecules that play key regulatory roles in plant development, organ morphogenesis, and stress response signaling. In this study, applying computational approaches and following high stringent filtering criteria, a total of 40 potential microRNAs belonging to 19 families were characterized from guava. The identified miRNA precursors formed stable stem-loop structures and exhibited high sequence conservation among diverse and evolutionarily distant plant species. Differential expression pattern of seven selected guava miRNAs (pgu-miR156f-5p, pgu-miR160c-5p, pgu-miR162-3p, pgu-miR164b-5p, pgu-miR166t, pgu-miR167a-5p, and pgu-miR390b-5p) were recorded under salin...
MicroRNAs (miRNAs) are a key gene regulator and play essential roles in several biological and pa... more MicroRNAs (miRNAs) are a key gene regulator and play essential roles in several biological and pathological mechanisms in the human system. In recent years, plenty of miRNAs have been identified to be involved in the development of neurodegenerative disorders (NDDs), thus making them an attractive option for therapeutic approaches. Hence, in this review, we provide an overview of the current research of miRNA-based therapeutics for a selected set of NDDs, either for their high prevalence or lethality, such as Alzheimer's, Parkinson's, Huntington's, Amyotrophic Lateral Sclerosis, Friedreich's Ataxia, Spinal Muscular Atrophy, and Frontotemporal Dementia. We also discuss the relevant delivery techniques, pertinent outcomes, their limitations, and their potential to become a new generation of human therapeutic drugs in the near future.
Superfruit guava (Psidium guajava L.) is one of the healthiest fruits due to its high antioxidant... more Superfruit guava (Psidium guajava L.) is one of the healthiest fruits due to its high antioxidant dietary fiber and vitamin content. However, the growth and development of this plant are severely affected by salinity stress, mostly at the seedling stage. MicroRNAs (miRNAs) are small, noncoding, endogenous, highly conserved RNA molecules that play key regulatory roles in plant development, organ morphogenesis, and stress response signaling. In this study, applying computational approaches and following high stringent filtering criteria, a total of 40 potential microRNAs belonging to 19 families were characterized from guava. The identified miRNA precursors formed stable stem-loop structures and exhibited high sequence conservation among diverse and evolutionarily distant plant species. Differential expression pattern of seven selected guava miRNAs (pgu-miR156f-5p, pgu-miR160c-5p, pgu-miR162-3p, pgu-miR164b-5p, pgu-miR166t, pgu-miR167a-5p, and pgu-miR390b-5p) were recorded under salinity stress and pgu-miR162-3p, pgu-miR164b-5p as well as pgu-miR166t were found to be the most affected ones. Using the psRNATarget tool, a total of 49 potential target transcripts of the characterized guava miRNAs were identified in this study which are mostly involved in metabolic pathways, cellular development, and stress response signaling. A biological network has also been constructed to understand the miRNA mediated gene regulation using the minimum free energy (MFE) values of the miRNA-target interaction. To the best of our knowledge, this is the first report of guava miRNAs and their targets.
Chikungunya (CHIKV) and Dengue (DENV) viruses cause an acute febrile illness which is hard to cli... more Chikungunya (CHIKV) and Dengue (DENV) viruses cause an acute febrile illness which is hard to clinically differentiate and treat since both exhibit similar symptoms. Hence, this study was aimed at identifying the expression profiles of cytokines on co-infected samples and compare with CHIKV and DENV mono-infected samples. Serum samples of 292 suspected patients during 2009-2011 were analyzed. The presence of primary (IgM)/secondary (IgG) antibodies and early NS1 Dengue antigens were confirmed by capture ELISA. Molecular diagnosis and serotypes were discriminated by RT-PCR, confirmed by sequencing. All the plasma samples were assayed for cytokine expression by BDTM cytometry bead array (CBA) and compared with independent mono-infection viral load. Among the tested samples, 82 were confirmed as Dengue positive; 52 through IgM (17.8%), and 30 through IgG (10.2%). Additionally, 186 samples were confirmed as Chikungunya, 96 through IgM (32.6%) and 92 through IgG (31.5%) ELISA, respectively. Interestingly , 19 patients were co-infection positive in which, only 6 were confirmed for CHIKV and 7 for DENV by RT-PCR. Among 8 cytokines, IL-2, IL-8, IFNα, IFN γ, and IL-12 were found to be significantly different between co-infected and CHIKV mono-infected patients and correlated with viral load. DENV viral load was correlated with cytokine expression and a significant difference in IL-2 and IL-12 was observed between DENV mono-infected and DENV and CHIKV co-infected patients. Results indicated that apart from serological and molecular confirmation, cytokines could be used as a specific biomarker for the diagnosis of DENV and CHIKV. In the future, the role of independent cytokines can be determined to understand the pathogenesis and etiology of these dreadful diseases.
MicroRNAs (miRNAs) are highly conserved, endogenous, short (21-24 nucleotides), non-coding RNA mo... more MicroRNAs (miRNAs) are highly conserved, endogenous, short (21-24 nucleotides), non-coding RNA molecules that play significant roles in post-transcriptional gene silencing by directing target mRNA cleavage or translational inhibition. Nonetheless, highly nutritious ´super grain´ quinoa (Chenopodium quinoa) is an extreme abiotic stress tolerant Andean seed crop of many potential uses with an outstanding protein quality and a load of vitamins, minerals as well as flavonoids antioxidant. In this study applying genome-wide in silico approaches (referring recently published quinoa genome) and following a set of stringent filtering measures a total of 22 potentially conserved microRNAs belonging to 18 families were characterized from quinoa and eleven randomly selected putative microRNAs (cqu-miR160a, cqu-miR162a, cqu-miR164a,
Background: The banana (Musa sp., AAA) genome is constantly increasing due to high-frequency of s... more Background: The banana (Musa sp., AAA) genome is constantly increasing due to high-frequency of somaclonal variations. Due to its large diversity, a conventional numerical and morphological based taxonomic identification of banana cultivars is laborious, difficult and often leads to subject of disagreements. Results: Hence, in the present study, we used universal DNA barcode ITS2 region to identify and to find the genetic relationship between the cultivars and varieties of banana. Herein, a total of 16 banana cultivars were PCR amplified using ITS2 primer pair. In addition, 321 sequences which were retrieved from GenBank, USA, were used in this study. The sequences were then aligned using Clustal W and genetic distances were computed using MEGA V5.1. The study showed significant divergence between the intra-and inter-specific genetic distances in ITS2 region. BLAST1 and Distance methods proved that ITS2 DNA barcode region successfully identified and distinguished the cultivar and varieties of banana.
Endophytic fungi represent a promising biotechnological tool to identify and produce in large sca... more Endophytic fungi represent a promising biotechnological tool to identify and produce in large scale novel anti-inflammatory bioactive compounds. In this study, Crescentia alata Kunth was selected following the eth-nomedical criteria and a total of 219 isolates grouped in 86 morphotypes were obtained. From these, 44 isolates that presented a pigment-producing morphotype were selected as the screening panel. The ITS2 ribotypes of the selected endophytic fungi were annotated and classified phylogenetically based on a sequence-structure analysis. The isolates belonged to 17 genera: Colletotrichum, Fusarium, Lophotrichus, Podo-spora, Xylaria, Diaporthe, Aspergillus, Periconia, Didymella, Prosthemium, Trematophoma, Cladosporium, Cerco-spora, Pseudocercosporella, Aureobasidium, Bjerkandera, and Trametes. From the anti-inflammatory screening with the Griess assay only 14.77% were highly active with no significant difference compared to indometha-cin, and showed promising in vitro anti-inflammatory effect tested in murine macrophages induced with bacterial lipopolysaccharides (LPS). Interestingly, 11.36% of the extracts increased the production of nitrite on LPS-induced macrophages. None of the extracts at the tested concentrations presented a pro-inflammatory effect on non-induced macrophages, nor a cytotoxic effect (cell viability >85%) in the resazurin bioassay. Metabolic profiling of the endo-metabolome and exo-metabolome extracts using Thin Layer Chromatography (TLC), revealed that the exo-metanolome extracts had a relative higher number and diversity of chemical groups. The 1 H NMR metabolomic analysis showed characteristic signals that differentiate the fungal genera with high anti-inflammatory activity from those with the least activity. These signals could be associated with the group of terpenes. This is the first report on the isolation of endophytes from C. alata, from which 13 isolates exhibit pharmacological value as sources of potential anti-inflammatory and immuno-modulatory compounds. These bioactive metabolites are likely to belong to the groups of terpenes.
Biomass, Biofuels, Biochemicals. https://doi.CHAPTER 4 actions. Such anticipated characteristics ... more Biomass, Biofuels, Biochemicals. https://doi.CHAPTER 4 actions. Such anticipated characteristics fulfill the highly requisite demands of sustainable development of robust biocatalysts following green chemistry agenda .
MDPI, 2020
Triple-negative breast cancer (TNBC) cells are deficient in estrogen, progesterone and ERBB2 rece... more Triple-negative breast cancer (TNBC) cells are deficient in estrogen, progesterone and ERBB2 receptor expression, presenting a particularly challenging therapeutic target due to their highly invasive nature and relatively low response to therapeutics. There is an absence of specific treatment strategies for this tumor subgroup, and hence TNBC is managed with conventional therapeutics, often leading to systemic relapse. In terms of histology and transcription profile these cancers have similarities to BRCA-1-linked breast cancers, and it is hypothesized that BRCA1 pathway is non-functional in this type of breast cancer. In this review article, we discuss the different receptors expressed by TNBC as well as the diversity of different signaling pathways targeted by TNBC therapeutics, for example, Notch, Hedgehog, Wnt/b-Catenin as well as TGF-beta signaling pathways. Additionally, many epidermal growth factor receptor (EGFR), poly (ADP-ribose) polymerase (PARP) and mammalian target of rapamycin (mTOR) inhibitors effectively inhibit the TNBCs, but they face challenges of either resistance to drugs or relapse. The resistance of TNBC to conventional therapeutic agents has helped in the advancement of advanced TNBC therapeutic approaches including hyperthermia, photodynamic therapy, as well as nanomedicine-based targeted therapeutics of drugs, miRNA, siRNA, and aptamers, which will also be discussed. Artificial intelligence is another tool that is presented to enhance the diagnosis of TNBC.
Molecules, 2018
Galphimia glauca (Cav.) Kuntze is an important endemic plant species, which possesses many medici... more Galphimia glauca (Cav.) Kuntze is an important endemic plant species, which possesses many medicinal properties and has been used in the Mexican traditional medicine for its sedative, anxiolytic, anticonvulsant, antiasthmatic and antiallergic properties. The therapeutic properties of this plant are mainly due to the presence of diverse bioactive compounds such as flavonoids, triterpenoids, and phenolics. Several triterpenoids and flavonoids compounds have been isolated and identified. Modern studies have demonstrated many biological activities such as anti-inflammatory, antidiarrheal, gastroenteritis, antimalarial and cytotoxic activities. Nevertheless, many studies are restricted to the crude extract, and many bioactive compounds are yet to be identified and validated according to its traditional use. However, its commercial exploitation and use are highly limited due to the non-availability of enough plant material and lack of knowledge about its agronomical practices. Moreover, the misinterpretation and mislabeling of closely related species of the genus Galphimia Cav. as G. glauca or G. gracilis is a common problem for its rigorous scientific study and commercial exploitation. The present review provides comprehensive knowledge based on the available scientific literature. To the best of our knowledge, this is the first review on G. glauca. This comprehensive information will certainly provide a guide for the better understanding and utilization of G. glauca for its scientific and industrial exploitation.
Journal of Natural products, 2019
Two new prenylated acylphloroglucinols, paleacenins A (1) and B (2), were isolated from the rhizo... more Two new prenylated acylphloroglucinols, paleacenins A (1) and B (2), were isolated from the rhizome n-hexane and chloroform extracts of the fern Elaphoglossum paleaceum. Both compounds were found to possess the same geranylated filicinic acid moiety but have a different phloroglucinol ring substituent. Their structures were determined using 1 H and 13 C NMR spectroscopic, HRMS, and ECD analysis. The plant extracts and purified compounds were assayed for inhibition of monoamine oxidase (MAO) activity, and the n-hexane and chloroform extracts displayed 25.0% and 26.5% inhibition of MAO-A, respectively, as well as 42.5% and 23.7% inhibition of MAO-B, respectively. Compounds 1 and 2 exhibited IC 50 values of 31.0 (1.3) μM for MAO-A and 4.7 (4.4) μM for MAO-B. Paleacenin A (1) showed a higher selective index (SI) toward MAO-B (SI MAO-B/MAO-A 0.1), and paleacenin B (2) exhibited selectivity to MAO-A (SI MAO-B/MAO-A , 3.5). The extracts showed cytotoxicity against a panel of prostate, cervix, breast, and colon cancer cell lines (IC 50 values between 1.7 and 10.6 μg/mL); the pure compounds were more active against the prostate, cervix, and colon cancer cell lines. Paleacenins A (1) and B (2), with IC 50 values of 46 and 41 μM, respectively, inhibited nitric oxide production by the RAW264.7 murine macrophage model.
Molecular Biology Report, 2019
Ovalbumin is considered a protein of high nutritional value because it contains essential amino a... more Ovalbumin is considered a protein of high nutritional value because it contains essential amino acids and is highly digestible. Therefore, it has a high biological value. Currently, the high food demand requires worldwide attention because food production is insufficient. Therefore, other alternatives are necessary to satisfy food demands, such as protein engineering. In this work, a protein with a high essential amino acid content similar to ovalbumin was synthesized by protein engineering , expressed, and digested in vitro. The assembly and sequential overlap extension PCR strategy was used to synthesize a 345-bp gene that encodes a high essential amino acid content protein (HEAAP). The 345-bp product was cloned into the vector pBAD TOPO®, and expressed in Escherichia coli BL21. PCR reactions and sequencing demonstrated the presence, orientation, and correct sequence of the insert. HEAAP expression was induced by l-arabinose and then purified using Ni-NTA affinity chromatography. The expression in E. coli was low and barely detected by Western blot assay. The in vitro multienzyme digestibility of HEAAP was around 79%, which suggests that the protein is potentially nutritious. Virtual analysis classifies the protein as unstable and hydrophilic, with a half-life in E. coli of 10 h. The recombinant HEAAP was successfully synthesized, but it is necessary to improve the digestibility and to optimize expression including selecting other expression models.
Journal of the Institute of Tropical Medicine of São Paulo, 2019
Multidrug resistance (MDR), virulence and transferable elements potentiate Pseudomonas aeruginosa... more Multidrug resistance (MDR), virulence and transferable elements potentiate Pseudomonas aeruginosa's role as an opportunistic pathogen creating a high risk for public health. In this study, we evaluated the possible association of multidrug resistance, virulence factors and integrons with intrahospital P. aeruginosa strains isolated from patients at Cumana hospital, Venezuela. Relevant clinical-epidemiological data were collected to study 176 strains (2009-2016) isolated from different hospital units. Bacterial resistance was classified as susceptible, low-level resistant (LDR), multidrug resistant (MDR) and extensively drug-resistant (XDR). Most strains produced pyoverdine, DNase, gelatinase and hemolysin. Around 73% of the strains showed some type of movement. MDR and XDR strains increased from 2009 (24.2% and 4.8%, respectively) to 2016 (53.1% and 18.8%); while LDR decreased from 64.5% to 6.3%. The exoU and exoS genes were found in a significant number of strains (38.1 and 7.4%, respectively). Class I integrons were detected in 35.8% of the strains and the frequency was associated with resistance (42.9, 22.4, 41.4 and 61.9%, for susceptible, LDR, MDR and XDR, respectively). The MDR/XDR strains were positively associated with hemolysins and exoU, but negatively associated with bacterial twitching. MDR/XDR phenotypes were also associated with the Intensive Care Unit (ICU), septicemia, bronchial infection and diabetic foot ulcers, as well as long hospital stay (≥10 days) and previous antimicrobial treatment. High frequency of MDR/XDR strains and their association with class I integrons and virulence factors can increase the infection potential, as well as morbidity and mortality of patients attending this hospital and could spread infection to the community, creating a health risk for the region.
3Biotech, 2019
In the present study, a novel transformation protocol for Opuntia ficus-indica was generated by m... more In the present study, a novel transformation protocol for Opuntia ficus-indica was generated by means of particle bombardment. The best conditions obtained were: 900 psi rupture disk pressure, 8 cm microprojectile travel distance, and 4 h of exposition to 0.2 M mannitol. For all experiments, gold particles coated with 1.0 µg/µL of pBI426 plasmid DNA were used. With all these conditions, a 23% of transformation efficiency in terms of regeneration in selection media (100 mg/L kanamy-cin) was obtained. Interestingly, the presence of both transgenes: nptII and uidA, by means of PCR and RT-PCR assays was detected. The regeneration percentage achieved in terms of stable integration for both genes was 10%. In addition, we also detected adequate amounts of β-glucuronidase activity by means of the GUS fluorometric assay. The procedure described in the present investigation reveals the feasibility of using nopal for the introduction, expression, and possible production of heterologous proteins.
Life, 2019
In recent years, metabolic engineering of microorganisms has attained much research interest to p... more In recent years, metabolic engineering of microorganisms has attained much research interest to produce biofuels and industrially pertinent chemicals. Owing to the relatively fast growth rate, genetic malleability, and carbon neutral production process, cyanobacteria has been recognized as a specialized microorganism with a significant biotechnological perspective. Metabolically engineering cyanobacterial strains have shown great potential for the photosynthetic production of an array of valuable native or non-native chemicals and metabolites with profound agricultural and pharmaceutical significance using CO 2 as a building block. In recent years, substantial improvements in developing and introducing novel and efficient genetic tools such as genome-scale modeling, high throughput omics analyses, synthetic/system biology tools, metabolic flux analysis and clustered regularly interspaced short palindromic repeats (CRISPR)-associated nuclease (CRISPR/cas) systems have been made for engineering cyanobacterial strains. Use of these tools and technologies has led to a greater understanding of the host metabolism, as well as endogenous and heterologous carbon regulation mechanisms which consequently results in the expansion of maximum productive ability and biochemical diversity. This review summarizes recent advances in engineering cyanobacteria to produce biofuel and industrially relevant fine chemicals of high interest. Moreover, the development and applications of cutting-edge toolboxes such as the CRISPR-cas9 system, synthetic biology, high-throughput "omics", and metabolic flux analysis to engineer cyanobacteria for large-scale cultivation are also discussed.
Phyton, 2019
Torin 1 is an ATP-competitive TOR inhibitor which inhibits the signaling of TOR and S6K kinase in... more Torin 1 is an ATP-competitive TOR inhibitor which inhibits the signaling of TOR and S6K kinase in mammals and plants. The objective of this research is to determine the effect of Torin 1 in a relatively simple and homogeneous plant system such as the NT-1 tobacco suspension cell cultures. Cultures of NT-1 cells were tested with 5, 50, 150 and 250 nM of Torin 1. During kinetics growth of NT-1 tobacco suspension cell cultures, 150 and 250 nM Torin 1 inhibits the early growth and later enhanced the cellular proliferation during exponential growth by means of an increased expression of E2F1 and cyclin B. Furthermore, Torin 1 stimulates the growth of NT-1 cells during log phase with small shaped cell, characteristic of tobacco suspension cell cultures with high mitotic activity.
Canadian Journal of Plant Sciences, 2019
MicroRNAs (miRNAs) are highly conserved, endogenous, short (21-24 nucleotides), non-coding RNA mo... more MicroRNAs (miRNAs) are highly conserved, endogenous, short (21-24 nucleotides), non-coding RNA molecules that play significant roles in post-transcriptional gene silencing by directing target mRNA cleavage or translational inhibition. Nonetheless, highly nutritious "super grain" quinoa (Chenopodium quinoa) is an extreme abiotic stress tolerant Andean seed crop of many potential uses, with outstanding protein quality and a load of vitamins, minerals, as well as flavonoid antioxidants. In this study, applying genome-wide in silico approaches (referring to the recently published quinoa genome) and following a set of stringent filtering measures, a total of 22 potentially conserved microRNAs belonging to 18 families were characterized from quinoa and 11 randomly selected putative microRNAs cqu-miR398b) were validated successfully by RT-PCR. Using the psRNATarget tool, a sum of 59 potential miRNA targets, mostly transcription factors, were identified that are involved in biosynthesis, metabolic processes, and signal transduction. Among the detected targets, six target transcripts (F-Box proteins, TCP, MYB, WD protein, NAC, and CSD) were reported to have specific roles in both flavonoids biosynthesis and stress response signaling in some plants. To the best of our knowledge, this is the first report of quinoa microRNAs and their targets. Résumé : Les microARN (miARN) sont de courtes molécules d'ARN (21 à 24 nucléotides) endogènes, fort bien conservées, qui ne codent rien, mais jouent un rôle important dans le silençage de l'expression génique après la transcription par le clivage de certains ARNm ou l'inhibition de leur traduction. Le quinoa (Chenopodium quinoa), une super céréale, est une culture grainière des Andes qui tolère extrêmement bien les stress abiotiques. Elle possède de nombreux usages potentiels et sa protéine, de qualité exceptionnelle, renferme une foule de vitamines, d'oligoéléments et de flavonoïdes antioxydants. Les auteurs ont appliqué une approche in silico au génome complet (en recourant au génome du quinoa récemment publié) et recouru à un jeu de mesures de filtrage rigoureuses pour recueillir 22 microARN éventuellement conservés appartenant à 18 familles du quinoa. Onze microARN pos-cqu-miR398b), ont été validés par RT-PCR. Grâce à l'outil psRNATarget, les chercheurs ont identifié 59 cibles potentielles intervenant dans la photosynthèse, divers proces-sus métaboliques et la transduction des signaux pour ces miRNA, essentiellement des facteurs de transcription. Parmi elles figurent six transcrits (protéines F-Box, TCP, MYB, WD, NAC, et CSD) qui jouent un rôle spécifique dans la biosynthèse des flavonoïdes et le signalement de la réaction au stress chez quelques plantes. Pour autant qu'on le sache, il s'agit de la première fois où l'on rapporte l'existence de microARN et de leurs cibles chez le quinoa. [Traduit par la Rédaction]
Applied Nanoscience, 2020
Copper oxide nanoparticles possess a high absorption coefficient and are non-toxic to animal cell... more Copper oxide nanoparticles possess a high absorption coefficient and are non-toxic to animal cells. Biological agents inhabit many compounds that can be explored for the synthesis of monodispersed and non-toxic copper oxide nanoparticles. This is the most important advantage of biological syntheses over physical and chemical methods. In the present work, copper oxide nanoparticles have been synthesized with a simple and green technique by using an aqueous extract of mixed leaf and flowers of Galphimia glauca. The nanoparticles were synthesized at 80 °C, with a 15 mM Copper sulfate solution and at varying pH range (2, 4, inherent, i.e. 5.3, 8, 10, 12). The particle size of the nanoparticles was confirmed to be 5-10 nm using TEM and XRD at pH 12. The functional groups on the surface of the nanoparticles were characterized using FTIR. Moreover, GCMS analysis of the aqueous extracts showed that different flavonoids were responsible for the biosynthesis of copper oxide nanoparticles. The cytotoxicity assays were also determined using MRC-5 and HeLa cell lines for copper oxide nanoparticles and was found that the nanoparticles are non-toxic to the normal cells and are relatively toxic to cancer cells.
Journal of Material Science, 2019
There are enormous methods such as physical, chemical, and biological, for the synthesis of metal... more There are enormous methods such as physical, chemical, and biological, for the synthesis of metallic nanoparticles (MNPs), which has become a matter of focus among material scientists. Green chemistry-based MNP synthesis is an area, which has gained much importance presently due to their non-toxicity and monodispersed nanoparticle preparation methodologies. Among green synthesis methods, plants are considered as efficient candidates for nanoparticle synthesis. The meticulous formation of different sizes and shapes of the nanoparticles using plants has spurred encouraging interest. The rate kinetics and stability of nanoparticle synthesis are well studied as well as appreciated in the arena of materials. Their capability to sequester metal ions and fastidiously define the dimensions using a plethora of capping proteins such as glutathione and phytochelatins is intriguing giving it a monodispersed size. This review is a comprehensive understanding of the metal nanoparticles synthesized by plants and apprehends the mechanism of nanoparticle synthesis exhaustively.
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Papers by Dr. Ashutosh Sharma
range of both early and late colonizers. Organism communities making up biofilms arrange themselves nonrandomly into
colonies of bacterial cells, yeasts, and other fungi, a matrix known as glycocalyx. Aerobic species occupy the biofilm
surface, whereas anaerobic species arrange themselves in deeper layers, which creates actual gas diffusion gradients. Typically,
biofilms begin their formation by propagating laterally, and after these first developmental stages, biofilm begins to
grow vertically (Fig. 5.1) (Socransky and Haffajee, 2002, 2005).
Thus, differential geographical locations are produced during the first growth phases, while dense accumulations of
microorganisms attached to organic or inorganic polysaccharide matrices take place in lower biofilm layers, where stationary
groups can be differentiated from free groups. Biofilms are dynamic communities where nutritional components
and gases are transported by molecular diffusion (Lang and Tonetti, 2003). Microbial colonies are often composed of different
species within a matrix surrounded by water channels that allow for nutrients and other agents to pass, resembling a
primitive circulatory system. These channels create an enormous maze consisting mostly of exopolysaccharides, whose
role is essential in safeguarding the integrity of a constantly-growing biofilm (Socransky and Haffajee, 2002). Within
the biofilm, at the microbial species level, cell physiology is quite heterogeneous, which means that cells can be at different
physiological stages even when they are close to one another. When environmental factors affect one species, cells of the
same species in deeper layers of the biofilm can remain unharmed (Marsh, 2005). The human host contributes directly, due
to lack of dental hygiene, or indirectly, by ingestion, to the promotion of biofilm growth and stability, making the oral cavity
a haven for microbial resistance (Marsh and Devine, 2011) (Fig. 5.2). The complexity of the cell-to-cell interactions
required for the biofilm formation and stability is worth the advantages of physical protection obtained by colonizers from
hostile factors in the oral environment. Most often, these factors are variations in pH, temperature, redox reactions, and
nutrient concentration, as well as potentially toxic substances such as antibiotics and the host’s immunological response
mechanisms (Socransky and Haffajee, 2002).