Temporal Temperature Gradient Research Articles

Consider Non-Syndromic Mitochondrial Disorders as Part of the Pediatric Phenotypic Spectrum

With interest we read the article by Ardissone et al. about a retrospective, singlecenter study of 150 pediatric patients with a genetically confirmed mitochondrial disorder (MID) due to a pathogenic mtDNA variant (mtDNA point mutation or single mtDNA deletion) [1]. The study is excellent but has limitations that are cause of concerns and should be discussed. A limitation of the study is that only syndromic MIDs were included [1]. Since MIDs often do not fit into any of the syndromic phenotypes, we should know whether these patients were deliberately excluded or did not appear at the study center at all. We should know whether the entire group of 902 patients also included patients with nonsyndromic MIDs. Another limitation is that heteroplasmy rates were only determined by estimating the densitometric ratio between wild-type and mutant mtDNA [1]. Heteroplasmy is usually determined by a number of quantitative methods, such as Sanger sequencing, high performance liquid chromatography, pyrosequencing, Snapshot, high resolution melt profiling (HRM), temporal temperature gradient gel electrophoresis (TTGE), invader assay, amplification refractory mutation system (ARMS), endonuclease method using Surveyor nuclease, and next generation sequencing (NGS) [2]. These techniques usually provide more accurate results than semi-quantitative estimation. There is a discrepancy between the statement in the method section ("only few data were not available from all patients”) and the results section (“CSF data were available from only 17 %, follow-up data only from 47%, and biochemical data only from 58 % of patients”) [1]. This discrepancy should be clarified. Another limitation is that the total observation time up to the last follow-up was not given [1]. Knowing this latency is important to assess the speed of disease progression and estimate the outcome. There is also a discrepancy between the statement in the result section that “outcome data were evaluated from 70 patients” and the detailed description in the results section, which included 71 patients. Which number is correct? Surprisingly, the biochemical examination was normal in almost a quarter of the patients with Leigh syndrome. We should know if these results are from studies of just muscle or fibroblasts, or if other tissues have been studied as well If only muscle or fibroblasts were examined, we should know whether or not these tissues were clinically affected in the one quarter of the Leigh patients. Surprisingly, only two thirds of the MELAS patients presented with stroke-like episodes (SLEs), which are pathognomonic for MELAS. We should know if this was due to the short observation or follow-up time in these patients. According to what criteria were these patients without SLE classified as MELAS? Were the Hirano or Japanese criteria applied? We should also know how “MELAS-like” was defined. There is no mention of the underlying mutations in the PEO patients. It would be interesting to know if these patients all carried a single mtDNA deletion, a nuclear DNA-located variant, or an mtDNA point mutation. Overall, the interesting study has limitations that call the results and their interpretation into question. With a prospective design and extensive and long-term follow-up, the study goals could be better achieved.

A hybrid approach to predict vertical temperature gradient of ballastless track caused by solar radiation

• A hybrid approach combining the field test and FEM with ML technique is proposed. • Six ML techniques are used to predict spatial and temporal temperature gradient. • The key features of hybrid approach to predict temperature gradient are analyzed. • The proposed hybrid approach has excellent performance and transferability. The spatial and temporal temperature evolution of ballastless track is difficult to be determined, due to its complex thermal conditions. Accurate acquisition of vertical temperature gradient (VTG) of ballastless track is of great significance for its design, maintenance and damage control. However, few researchers have studied the predictive approach of spatial and temporal VTG evolution. In this study, a hybrid approach that combines experimental and finite element analysis with machine learning (ML) technique is proposed to automatically predict VTG of ballastless track induced by solar radiation. Six different ML techniques, i.e., multiple linear regression (MLR), polynomial regression (PR), support vector regression (SVR), random forest (RF), extreme gradient boosting (XGBOOST), and categorical boosting (CATBOOST) with fine-tuned hyperparameters are trained and validated by a dataset of 1000 cases retrieved from macroscale finite element analysis. The maximum and minimum air temperatures, latitude, average wind speed, etc., considering spatial and temporal environment actions are used as input variables to estimate the VTG of ballastless track. The results indicate that the CATBOOST-based hybrid approach demonstrates the most appropriate method for determining VTG of ballastless track among the above ML techniques. The proposed feasible and efficient hybrid approach is expected to be widely used for the identification of temperature evolution in structural engineering.

Effect of daily co-exposure to inulin and chlorpyrifos on selected microbiota endpoints in the SHIME® model

The intestinal microbiota has a key role in human health via the interaction with the somatic and immune cells in the digestive tract environment. Food, through matrix effect, nutrient and non-nutrient molecules, is a key regulator of microbiota diversity. As a food contaminant, the pesticide chlorpyrifos (CPF) has an effect on the composition of the intestinal microbiota and induces perturbation of microbiota. Prebiotics (and notably inulin) are known for their ability to promote an equilibrium of the microbiota that favours saccharolytic bacteria. The SHIME® dynamic in vitro model of the human intestine was exposed to CPF and inulin concomitantly for 30 days, in order to assess variations in both the bacterial populations and their metabolites. Various analyses of the microbiota (notably temporal temperature gradient gel electrophoresis) revealed a protective effect of the prebiotic through inhibition of the enterobacterial (E. coli) population. Bifidobacteria were only temporarily inhibited at D15 and recovered at D30. Although other potentially beneficial populations (lactobacilli) were not greatly modified, their activity and that of the saccharolytic bacteria in general were highlighted by an increase in levels of short-chain fatty acids and more specifically butyrate. Given the known role of host-microbiota communication, CPF's impact on the body's homeostasis remains to be determined.

Pyroelectric performance of [Bi0.48Na0.4032K0.0768]Sr0.04(Ti0.975Nb0.025)O3 ceramics

In the present work, the pyroelectric performance of [Bi0.48Na0.4032K0.0768]Sr0.04(Ti0.975Nb0.025)O3 (BNT-2.5Nb) ceramics is investigated. At room temperature, the value of pyroelectric coefficient found as 13.2 × 10−4 C/m2 K which is higher than many lead-free ferroelectric materials. Further, the pyroelectric figures of merit (FOMs) for detectivity (Fd), voltage responsivity (Fv), current responsivity (Fi), and energy harvesting (Fe*) are calculated. BNT-2.5Nb ceramics show pyroelectric open circuit voltage as 0.45 V when it was exposed to temporal temperature gradient. This work result indicates that the Nb-doping in Bi0.5Na0.5TiO3-based composition can be beneficial for lead-free pyroelectric device applications.

Diesel Exhaust Emission Soot Coated Pyroelectric Materials for Improved Thermal Energy Harvesting.

Pyroelectric performance is significantly improved using a coating of diesel exhaust soot. Coated pyroelectric sample (lead zirconate titanate) is exposed to a temporal temperature gradient. Under the application of infrared (IR) heating for a given temperature gradient, the maximum open circuit voltage improves more than four times, electric current across a resistance of 10 ohm improves more than six times, and the stored energy in 10 µF capacitor is enhanced by 17 times. These results are important from two aspects: 1) utilization of waste diesel exhaust soot and 2) improving energy harvesting performance of pyroelectric materials.

Estructura de la comunidad bacteriana en diferentes tejidos de Lobatus gigas silvestres (Linnaeus, 1758) de la Reserva de Biosfera Seaflower del Caribe

The microbial diversity of Lobatus gigas has not been thoroughly studied despite of them is a specie endangered. Knowledge of microbiota may help to improve the conservation and cultivation of this species. The objective of this study was to evaluate the bacterial populationsassociated with the gonad and the gut compartments of the wild endangered L. gigas from the Caribbean Seaflower Biosphere Reserve, using microbiological methods and culture-independent molecular tools. The genetic profiles of the bacterial populations were generated and Temporal Temperature Gradient Electrophoresis (TTGE) was used to compare them with total DNA. A genetic and statistical analysis of the bacterial communities revealed a low level of diversity in gonad tissue based on the number of bands detected using TTGE. In addition, statistical differences in bacterial community structure were found between the foregut and hindgut tissue of L. gigas. The dominant phylogenetic affiliations of the gonad bacteria, as determined using 16S rRNA gene sequencing, belong to Ralstonia (50%). The possible involvement of this genus in the reproduction and development of the conch is discussed. On the other hand, the bacterial phylotypes from foregut and hindgut included members of Alphaproteobactera (12.5%), Betaproteobacteria (12.5%), Gammaproteobacteria (12.5%), Bacilli (31.25%), Clostridia (6.25%), Actinobacteria (6.25%), Mollicutes (6.25%) and Deinococci (6.25%) classes. Knowing the composition of the gonad and foregut and hindgut bacteria of L. gigas is the first step toward exploring the proper management of this species, as well as provides useful information to future researches that allow a better understanding of the role of these bacterial populations in the health and reproductive rate of L. gigas.

High bacterial activity in nutrient rich saltwater: Evidence from the uncoupling between salinity and nutrients in the Patos Lagoon estuary

In estuaries, a common pattern found is of higher bacterial activity in freshwater than in saltwater, an indication of the existence of an activity gradient. It is not clear, however, whether this arises from a natural property of saltwater bacterial communities or from metabolic constraints imposed from the natural co-variation between salinity and nutrients in estuarine systems. In this study, we managed to detect in situ higher activity levels of free-living bacteria in nutrient-rich saltwater entering the Patos Lagoon estuary. The bacterial community composition (BCC - Temporal Temperature Gradient Gel Electrophoresis) and activity (community-level physiological profiles, CLPP- Ecoplate™ - Biolog) were evaluated during two days in a transect along the estuary until the adjacent coastal region. The unexpected higher nutrient content and bacterial activity in saltwater found suggest that the bacterial activity is a primary response to nutrient availability, whereas salinity is the main factor structuring BCC in this estuary. Our results reject the idea of a natural low activity of saltwater bacteria by showing that their activity can increase in situ given that the proper conditions are met, especially in estuaries that display circulation patterns that promote the occurrence of nutrient-rich saltwater.

Pyroelectric performance of [(1-x)Ba0.9Ca0.1TiO3-x(BaSn0.2Ti0.8O3)] lead free ceramics

ABSTRACTIn the current work, the pyroelectric properties of [(1-x)Ba0.9Ca0.1TiO3-x(BaSn0.2Ti0.8O3)] lead free ceramics with x = 0, 0.4, 0.5 and 0.6 were investigated. The pyroelectric performance was found to increase with increase in the value of x. Pyroelectric analysis reveals an open circuit voltage of 0.52, 0.67, 0.78 and 0.90 V and also voltage at load 12 MΩ of 0.29, 0.35, 0.42 and 0.48 V for x = 0, 0.4, 0.5 and 0.6 respectively when it was exposed under temporal temperature gradient. The results show that [(1-x)Ba0.9Ca0.1TiO3-x(BaSn0.2Ti0.8O3)] (for x = 0.6) will be one of the potential material for pyroelectric applications.

Pyroelectric performance of BaTi1‐xSnxO3 ceramics

AbstractIn the present work, the pyroelectric properties of Sn‐doped BaTiO3 (BaTi1‐xSnxO3) were investigated. A significant increase in pyroelectric coefficient was observed for x = 0.05 composition and found to be 10 × 10−4 C/m2 K at 350 K. Further, the pyroelectric figure of merits (FOMs) for current responsivity (Fi), voltage responsivity (Fv), and energy harvesting (Fe and ) were calculated for all investigated samples. A large improvement in FOMs was achieved for x = 0.05 compared with undoped ceramic. BaTi1‐xSnxO3 (x = 0.05) was subjected to further pyroelectric analysis revealing an open‐circuit voltage of 0.6 V when it was exposed to temporal temperature gradient. The voltage and current were also measured across resistances 100 kΩ, 1 MΩ, and 5 MΩ. The results reveal that BaTi0.95Sn0.05O3 will be one of the potential lead‐free pyroelectric materials.

Effect of Temperature and Precipitation on Global Mangrove <i>Rhizophora</i> Species Distribution

Climate change influences the distribution of mangroves towards tropical or temperate regions of the world. We hypothesized that Rhizophora species organized along complex spatial and temporal temperature and precipitation gradient. To determine the affinity of different mangroves with climatic parameters, the response of eight Rhizophora species to seven climatic parameters (i.e., annual mean temperature, annual precipitation, maximum temperature of warmest month, minimum temperature of coldest month, temperature annual range and temperature of driest month and precipitation of warmest month) was analyzed with Arc GIS and Maxent software. For temperature and precipitation there was significant difference (p<0.001) among species location. Only R. selala (p<0.001) differed most from the rest taxa in temperature. The jackknife test in Maxent revealed that minimum temperature of coldest month (regularized training gain ~0.71), temperature annual range, (regularized training gain ~0.66) and annual mean temperature (regularized training gain ~0.36) were the most important environmental parameters that determine the occurrence of R. mangle, being the most dominant species. This outcome was further validated by an accuracy of 91%. This implies thattemperature was a better predictive parameter than precipitation in the distribution of R. mangle.

Characterization of lactic acid bacterial communities associated with a traditional Colombian cheese: Double cream cheese

Culture-dependent and Independent methods such as temporal temperature gradient electrophoresis (TTGE) and evaluation of the technological properties of five strains representing isolated groups, were used to evaluate the bacterial composition of three traditional double cream cheese companies in Colombia-South America. The results showed that fat, protein, moisture, and titratable acidity were significantly different between cheeses. Genetic analysis of these bacterial communities revealed a low level of diversity, based on the TTGE analysis. The dominant phylogenetic affiliations were grouped into four genera Enterococcus, Lactobacillus, Leuconostoc, and Lactococcus, related to acidification processes and formation of aromas and flavours in the cheese. The characterization of lactic acid bacteria (LAB) isolates showed predominance of the genera Leuconostoc and Weissella. These bacteria are related to the development of certain features such as texture and production of antimicrobial metabolites. In the evaluation of technological properties Weissella viridescens and Weissella paramesenteroides displayed greater pH reduction at 24 h, Pediococcus acidilactici, Leuconostoc mesenteroides and Leuconostoc citreum presented diacetyl production. The combination of different approaches allowed inferring the presence of a dominant population of LAB present in this cheese, this results complements the information needed for the establishment of a product protected under the status of Designation of Origin.

First Report of Swim Bladder-Associated Microbiota in Rainbow Trout (Oncorhynchus mykiss).

The aim of the present study was to identify major bacteria associated with the swim bladder in the rainbow trout, Oncorhynchus mykiss. We extracted DNA from the swim bladder and gut contents in order to perform a temporal temperature gradient gel electrophoresis (TTGE) analysis of 16S rRNA amplicons for bacterial identification to further compare both profiles. Arthrobacter and Cellulosimicrobium were the major genera observed in the swim bladder in fish, but were not present in fish gut contents; Mycoplasma were instead observed in these samples. Further research to investigate the possible symbiotic roles of the swim bladder-associated microbiota in salmonids is needed.

Diversity of the Bacterial Microbiota of Anopheles Mosquitoes from Binh Phuoc Province, Vietnam.

The naturally acquired microbiota of Anopheles can influence vector’s susceptibility to Plasmodium and its capacity to transmit them. Microbiota modification is a new challenge to limit disease transmission but it still needs advanced knowledges on bacterial community in Anopheles, especially in wild and infected specimens from diverse origin and species. Bacterial culture and 16S rRNA gene-PCR associated to Temporal Temperature Gradient Electrophoresis (TTGE) were applied to explore the bacterial diversity in the abdomen of 100 wild specimens (eight Anopheles species) collected in the Binh Phuoc Province, Vietnam. Culture and PCR-TTGE were complementary. The bacterial richness of the mosquito collection encompassed 105 genera belonging to seven phyla, mostly Proteobacteria, Firmicutes, and Actinobacteria. Staphylococcus, Clostridium, and Bacillus in Firmicutes were the most prevalent genera. However, Proteobacteria represented by 57 genera was the most diversified phylum in Anopheles microbiota. The high overall of Anopheles-associated bacteria is confirmed with, to our knowledge, 51 genera described for the first time in Anopheles microbiota. However, the diversity per specimen was low with average diversity index and the average Shannon–Wiener score (H) of 4.843 and 5.569, respectively. The most represented bacterial genera were present in <30% of the specimens. Consequently, the core microbiota share by Anopheles from Binh Phuoc was very narrow, suggesting that Anopheles microbiota was greatly influenced by local environments. The repertory of bacterial genera in two specimens of An. dirus and An. pampanai naturally infected by Plasmodium vivax was also described as preliminary results. Finally, this study completed the repertory of bacteria associated to wild Anopheles. Anopheles associated-bacteria appeared specimen-dependent rather than mosquitoe species- or group-dependent. Their origin and the existence of Anopheles-specific bacterial taxa are discussed.

Establishment and development of the intestinal microbiota of preterm infants in a Lebanese tertiary hospital

The establishment and development of the intestinal microbiota is known to be associated with profound short- and long-term effects on the health of full-term infants (FTI), but studies are just starting for preterm infants (PTI). The data also mostly come from western countries and little information is available for the Middle East. Here, we determined the composition and dynamics of the intestinal microbiota during the first month of life for PTI (n = 66) and FTI (n = 17) in Lebanon. Fecal samples were collected weekly and analyzed by quantitative PCR (q-PCR) and temporal temperature gradient gel electrophoresis (TTGE). We observed differences in the establishment and composition of the intestinal microbiota between the two groups. q-PCR showed that PTI were more highly colonized by Staphylococcus than FTI in the first three weeks of life; whereas FTI were more highly colonized by Clostridium clusters I and XI. At one month of life, PTI were mainly colonized by facultative anaerobes and a few strict anaerobes, such as Clostridium cluster I and Bifidobacterium. The type of feeding and antibiotic treatments significantly affected intestinal colonization. TTGE revealed low species diversity in both groups and high inter-individual variability in PTI. Our findings show that PTI had altered intestinal colonization with a higher occurrence of potential pathogens (Enterobacter, Clostridium sp) than FTI. This suggests the need for intervention strategies for PTI to modulate their intestinal microbiota and promote their health.

Gut microbiota related to Giardia duodenalis, Entamoeba spp. and Blastocystis hominis infections in humans from Côte d'Ivoire.

Literature data provide little information about protozoa infections and gut microbiota compositional shifts in humans. This preliminary study aimed to describe the fecal bacterial community composition of people from Côte d'Ivoire harboring Giardia duodenalis, Entamoeba spp., and Blastocystis hominis, in trying to discover possible alterations in their fecal microbiota structure related to the presence of such parasites. Twenty fecal samples were collected from people inhabiting three different localities of Côte d'Ivoire for copromicroscopic analysis and molecular identification of G. duodenalis, Entamoeba spp., and B. hominis. Temporal temperature gradient gel electrophoresis (TTGE) was used to obtain a fingerprint of the overall bacterial community; quantitative polymerase chain reaction (qPCR) was used to define the relative abundances of selected bacterial species/group, and multivariate statistical analyses were employed to correlate all data. Cluster analysis revealed a significant separation of TTGE profiles into four clusters (p < 0.0001), with a marked difference for G. duodenalis-positive samples in relation to the others (p = 5.4×10-6). Interestingly, qPCR data showed how G. duodenalis-positive samples were related to a dysbiotic condition that favors potentially harmful species (such as Escherichia coli), while Entamoeba spp./B. hominis-positive subjects were linked to a eubiotic condition, as shown by a significantly higher Faecalibacterium prausnitzii-Escherichia coli ratio. This preliminary investigation demonstrates a differential fecal microbiota structure in subjects infected with G. duodenalis or Entamoeba spp./B. hominis, paving the way for using further next-generation DNA technologies to better understand host-parasite-bacteria interactions, aimed at identifying potential indicators of microbiota changes.

Structural differences in gut bacteria communities in developmental stages of natural populations of Lutzomyia evansi from Colombia's Caribbean coast.

BackgroundLutzomyia evansi, a phlebotomine insect endemic to Colombia’s Caribbean coast, is considered to be the main vector of visceral and cutaneous leishmaniasis in the region. Although insects of this species can harbor pathogenic and non-pathogenic microorganisms in their intestinal microbiota, there is little information available about the diversity of gut bacteria present in Lutzomyia evansi. In this study, conventional microbiological methods and molecular tools were used to assess the composition of bacterial communities associated with Lutzomyia evansi guts in immature and adult stages of natural populations from the department of Sucre (Caribbean coast of Colombia).MethodsSand flies were collected from two locations (peri-urban and jungle biotype) in the Department of Sucre (Caribbean coast of Colombia). A total of 752 Lutzomyia evansi intestines were dissected. In this study, 125 bacterial strains were isolated from different culture media (LB Agar, MacConkey Agar). Different methods were used for bacterial identification, including ribosomal intergenic spacer analysis (RISA) and analysis of the 16S rRNA and gyrB gene sequences. The genetic profiles of the bacterial populations were generated and temporal temperature gradient gel electrophoresis (TTGE) was used to compare them with total gut DNA. We also used PCR and DNA sequence analysis to determine the presence of Wolbachia endosymbiont bacteria and Leishmania parasites.ResultsThe culture-dependent technique showed that the dominant intestinal bacteria isolated belong to Acinetobacter, Enterobacter, Pseudomonas, Ochrobactrum, Shinella and Paenibacillus in the larval stage; Lysobacter, Microbacterium, Streptomyces, Bacillus and Rummeliibacillus in the pupal stage; and Staphylococcus, Streptomyces, Brevibacterium, Acinetobacter, Enterobacter and Pantoea in the adult stage. Statistical analysis revealed significant differences between the fingerprint patterns of the PCR-TTGE bands in bacterial communities from immature and adult stages. Additionally, differences were found in bacterial community structure in fed females, unfed females, males and larvae. The intestinal bacteria detected by PCR-TTGE were Enterobacter cloacae and Bacillus thuringiensis, which were present in different life stages of Lu. evansi, and Burkholderia cenocepacia and Bacillus gibsonii, which were detected only in the larval stage. Wolbachia and Leishmania were not detected in gut samples of Lutzomyia evansi.ConclusionsThe analyses conducted using microbiological and molecular approaches indicated significant variations in the bacterial communities associated with the gut of Lu. evansi, depending on the developmental stage and food source. We propose that these elements affect microbial diversity in L. evansi guts and may in turn influence pathogen transmission to humans bitten by this insect.

WE-FG-202-01: Early Prediction of Radiotherapy Induced Skin Reactions Using Dynamic Infrared Imaging

Purpose: To predict radiotherapy induced skin reactions using dynamic infrared imaging. Methods: Thermal images were captured by our homebuilt system consisting of two flash lamps and an infrared (IR) camera. The surface temperature of the skin was first raised by ∼ 6 oC from ∼1 ms flashes. The camera then captured a series of IR images for 10 seconds. For each image, a baseline skin temperature was recorded for 0.5sec before heat impulse. The temporal temperature gradients were calculated between a reference point (immediately after the flash) and at a time point 9sec after that. Thermal effusivity, an intrinsic thermal property of a material, was calculated from the surface temperature decay of skin. We present experimental data in five patients undergoing radiation therapy, of which 2 were Head & Neck, 1 was Sarcoma and 2 were Breast cancer patients. The prescribed doses were 45 – 60 Gy in 25 – 30 fractions. Each patient was imaged before treatment and after every fifth fraction until end of the treatment course. An area on the skin, outside the radiation field, was imaged as control region. During imaging, each patient's irradiated skins were scored based on RTOG skin morbidity scoring criteria. Results: Temperature gradient, which is the temperature recovery rate, depends on the thermal properties of underlying tissue. It was observed that, the skin temperature and temporal temperature gradient increases with delivered radiation dose and skin RTOG score. The treatment does not change effusivity of superficial skin layer, however there was a significant difference in effusivity between treated and control areas at depth of ∼ 1.5 – 1.8 mm, increases with dose. Conclusion: The higher temporal temperature gradient and effusivity from irradiated areas suggest that there is more fluid under the irradiated skin, which causes faster temperature recovery. The mentioned effects may be predictors of Moist Desquamation.

Electrical Energy Generation from Hot/Cold Air Using Pyroelectric Ceramics

Low grade waste thermal energy is a prominent energy source for miniature devices. Present study reports experimental observations on thermal energy harvesting using pyroelectric ceramics (PbZrxTi1-xO3 (PZT-5H)). Appropriate electrical circuit was connected to the pyroelectric material for harvesting thermal energy. Hot/cold air was used to generate the continuous temporal temperature gradient. The temporal temperature variation was maintained as 4oC/sec. In order to study the behaviour of pyroelectric material for energy harvesting, generated charge was stored in capacitor. The maximum energy density was found to be 187.23 μJ/cm3 across 10 μF. Peak value of power output was 0.26 μW across 5MΩ and 4.7 μF.

Comparative Analysis of Denaturing Gradient Gel Electrophoresis and Temporal Temperature Gradient Gel Electrophoresis Profiles as a Tool for the Differentiation of Candida Species.

Background:Candida species are usually opportunistic organisms that cause acute to chronic infections when conditions in the host are favorable. Accurate identification of Candida species is an essential pre-requisite for improved therapeutic strategy. Identification of Candida species by conventional methods is time-consuming with low sensitivity, yet molecular approaches have provided an alternative way for early diagnosis of invasive candidiasis. Denaturing gradient gel electrophoresis (DGGE) and temporal temperature gradient gel electrophoresis (TTGE) are polymerase chain reaction (PCR)-based approaches that are used for studying the community structure of microorganisms. By using these methods, simultaneous identification of multiple yeast species will be possible and reliable results will be obtained quickly.Objectives:In this study, DGGE and TTGE methods were set up and evaluated for the detection of different Candida species, and their results were compared.Materials and Methods:Five different Candida species were cultured on potato dextrose agar medium for 24 hours. Next, total DNA was extracted by the phenol-chloroform method. Two sets of primers, ITS3-GC/ITS4 and NL1-GC/LS2 were applied to amplify the desired regions. The amplified fragments were then used to analyze DGGE and TTGE profiles.Results:The results showed that NL1-GC/LS2 primer set could yield species-specific amplicons, which were well distinguished and allowed better species discrimination than that generated by the ITS3-GC/ITS4 primer set, in both DGGE and TTGE profiles. All five Candida species were discriminated by DGGE and TTGE using the NL1-GC/LS2 primer set.Conclusions:Comparison of DGGE and TTGE profiles obtained from NL1-GC/LS2 amplicons exhibited the same patterns. Although both DGGE and TTGE techniques are capable of detecting Candida species, TTGE is recommended because of easier performance and lower costs.

Characterization of bacterial diversity associated with calcareous deposits and drip-waters, and isolation of calcifying bacteria from two Colombian mines

Bacterial carbonate precipitation has implications in geological processes and important biotechnological applications. Bacteria capable of precipitating carbonates have been isolated from different calcium carbonate deposits (speleothems) in caves, soil, freshwater and seawater around the world. However, the diversity of bacteria from calcareous deposits in Colombia, and their ability to precipitate carbonates, remains unknown. In this study, conventional microbiological methods and molecular tools, such as temporal temperature gradient electrophoresis (TTGE), were used to assess the composition of bacterial communities associated with carbonate deposits and drip-waters from two Colombian mines. A genetic analysis of these bacterial communities revealed a similar level of diversity, based on the number of bands detected using TTGE. The dominant phylogenetic affiliations of the bacteria, determined using 16S rRNA gene sequencing, were grouped into two phyla: Proteobacteria and Firmicutes. Within these phyla, seven genera were capable of precipitating calcium carbonates: Lysinibacillus, Bacillus, Strenotophomonas, Brevibacillus, Methylobacterium, Aeromicrobium and Acinetobacter. FTIR and SEM/EDX were used to analyze calcium carbonate crystals produced by isolated Acinetobacter gyllenbergii. The results showed that rhombohedral and angular calcite crystals with sizes of 90μm were precipitated. This research provides information regarding the presence of complex bacterial communities in secondary carbonate deposits from mines and their ability to precipitate calcium carbonate from calcareous deposits of Colombian mines.