Duration Of Incubation Research Articles

Antibiotic-Producing Streptomyces sp. Isolated from the Soil of a Mangrove Ecosystem

Highlights: 1. The unexplored soil of mangrove ecosystems in Surabaya, Indonesia, has the potential to be home to biodiversity,including Streptomyces sp. that can produce antibiotics.2. Streptomyces sp. has antibacterial properties against Gram-positive and Gram-negative bacteria, and the duration ofincubation plays a critical role in regulating the antibacterial activity. Abstract A mangrove ecosystem in Surabaya, Indonesia, has a high salinity, pH, potassium, phosphorus, and nitrate contents. This ecosystem comprises a mixture of sand, dust, mud, and clay, which has the potential to be a conducive environment for the isolation of Streptomyces. The importance of Streptomyces in biotechnology lies in its ability to produce bioactive secondary metabolites, which represent a valuable reservoir of antibiotics. This research aimed to assess the antibiotic activity exhibited by Streptomyces sp. isolated from the soil of a mangrove ecosystem in Wonorejo, Surabaya, Indonesia. The analysis focused on the potential of Streptomyces sp. to produce antibiotics that work against Gram-positive bacteria (i.e., Staphylococcus aureus ATCC 25923 and Bacillus subtilis) as well as Gram-negative bacteria (i.e., Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853, and Salmonella Typhimurium). The antibacterial activity test was conducted using the modified agar diffusion method. Observations were performed to identify any clear zone formation around the Streptomyces sp. agar colonies with a diameter of 0.8 cm and a height of 3 mm. The clear zone diameter was measured every 24 hours during the 10-day incubation period to assess the diversity of antibacterial activity. The antibacterial profile of Streptomyces sp. exhibited varying levels of activity against different bacterial strains in the tests conducted. The inhibition zone diameters demonstrated the highest levels of activity in Bacillus subtilis (15.9 mm) on day 7, Staphylococcus aureus (27.6 mm) on day 2, Pseudomonas aeruginosa (24.3 mm) on day 7, Escherichia coli (29.2 mm) on day 5, and Salmonella Typhimurium (27.5 mm) on day 7. The results indicated that Streptomyces sp. had inhibitory effects against Gram-positive bacteria as well as Gram-negative bacteria. In conclusion, Streptomyces sp. is a source of biodiversity found in the soil of mangrove ecosystems and has the ability to produce antibiotics.

Year-Round Spawning, Filial Cannibalism, and Embryonic and Larval Development of the Coral Reef Fish Orchid Dottyback, Pseudochromis fridmani

Aquaculture of marine ornamental fish could potentially reduce the fishing pressure on wild stocks by the global aquarium trade, but its expansion is often constrained by the limited understanding on the biology and early life history of candidate species. The orchid dottyback Pseudochromis fridmani is a valuable and popular marine ornamental fish, but scientific reports on its baseline biology, especially the egg filial cannibalistic behaviour, are scarce. The present study documented key aspects of reproductive biology and early ontogeny of P. fridmani in captivity, including reproductive behaviour, patterns of spawning and filial cannibalism throughout a 12-month period by seven pairs of brood fish, as well as the embryonic and larval development. The results showed that the captive spawning of the broodstock pairs generally occurred every 5–11 days, most commonly every 6–8 days, throughout a year. Despite feeding the brood fish to satiation throughout the data collection period, the average monthly rate of egg filial cannibalism reached 55 ± 37%, but the cannibalism frequency appeared to be pair-specific. The egg incubation duration was approximately 96 h at 27 ± 1 °C, and the development of embryos from cleavage, blastula, gastrula, and segmentation to the pharyngula stage is herein described in detail. The newly hatched P. fridmani larvae possessed important structures and organs for first feeding, including pigmented eyes, developed jaws, and a straight-tube gut. The 0 to 14 days post-hatching (DPH) period appeared to be an important larval stage, as P. fridmani larvae were observed to complete major changes in morphology, gut development, and phototactic behaviour by 14 DPH. Under the culture conditions of this study, the earliest transition to the juvenile stage was observed on 31 DPH, and the majority of fish became juveniles by 56 DPH. The results of this study inform baseline aquaculture production protocols and direct future research, particularly to reduce filial cannibalism through broodstock management and to improve larval culture through supporting the early ontogenetic development of P. fridmani. Additionally, these findings form a foundation for further studying the biology and ecology of P. fridmani in the wild.

Optimization of the Use of Industrial Wastes in Anaerobic Soil Disinfestation for the Control of Fusarium Wilt in Strawberry.

Anaerobic soil disinfestation (ASD) is proposed as an alternative to the use of chemical fumigants against Fusarium wilt in strawberry crops. Different residual wastes (rice bran, fishmeal, and residual strawberry extrudate) were assessed as amendments for ASD. Two different concentrations and two incubation durations were tested in growth chamber trials. The abundance of several microbial groups was noted before and after the treatments. Strawberry plants were grown in the treated soils to record Fusarium wilt disease severity. The population density of F. oxysporum increased after ASD in most amendments with rice bran and residual strawberry extrudate. Changes in Trichoderma spp., copiotrophic bacteria, and Streptomyces spp. populations were observed after anaerobiosis treatments and plant trials. A reduction in the disease severity was achieved in ASD-treated soils with 20 t/ha of rice bran at both 25 and 60 days of incubation, but not when using a 13.5 t/ha dose. Similarly, treatments using 19.3 t/ha of fishmeal for both incubation durations were able to reduce disease severity. In contrast, a severity reduction was only obtained in soils treated with 25.02 t/ha of the residual strawberry extrudate and incubated for 60 days in anaerobic conditions. Two of the three by-products tested were able to reduce Fusarium wilt symptoms in strawberry plants after an ASD-treatment period of only 25 days. Accordingly, the technique seems promising for strawberry growers in Huelva, Spain, and highly sustainable by giving value to residues produced in surrounding areas.

Abstract 19 CD34+ Cell Isolation from Cord Blood: A Novel Option for Faster, More Consistent Outcomes

Abstract Introduction Immunomagnetic isolation of cells has been long established but still suffers from drawbacks when used for hemopoietic stem cells (HSCs) in cord blood. [Kekarainen et al., 2006] Limitations with HSC isolation from cord blood include steep learning curves, time consuming manual labor, and inconsistent outcomes. [de Wynter et al., 1999; Perdomo-Arciniegas and Vernot, 2011] FerroBioTM has developed a semi-automated system for CD34+ cell isolation from unmodified cord blood (Fig 1A) as a means to provide fast, effective, and consistent isolations. Objectives The purpose of this work is to explore process improvements for the FerroBioTM system. Additionally, this work aims to characterize the resulting cell isolates. Methods The cell isolation protocol was evaluated by flow cytometry for cell counts, viability, and purity (total n=195). Protocol modification included incubation time at various steps, magnetic bead dosing, temperatures, and volumes. The ‘percent of cells captured’ was calculated as the difference between starting CD34 and non-captured CD34 cells. Cryopreservation tolerance was evaluated by freeze-thawing cells in a dimethyl sulfoxide-based media and measuring viability via hemocytometry (n=3). Sterility testing on cell isolates was performed via BacT/ALERT iNST & iAST (n=2). Results Incubation time did not have a significant effect on the efficiency of bead-cell binding (Fig 1B). As the FerroBioTM protocol progressed, final cell purity and viability increased and became more consistent (Fig 1C, D). Additionally, over the same phases of process development, final cell recovery increased 4-fold, from 6.5%±5.9% to 28.1%±9.7%. Post-cryopreservation viability was 78.8%±3.3%. Sterility tests were negative for anaerobic and aerobic bacteria. Discussion By establishing that bead-cell incubation duration did not significantly influence outcomes, the incubation time was considerably reduced, bringing the total processing time to ~3hrs. That said, if an overnight incubation is preferable for an operator’s workflow, the data suggests the process could be flexible to meet such needs. Iterations of the process have also resulted in significant improvements to final cell recovery while maintaining high purity and viability. Additional work is being done to further improve recovery. The initial investigation of sterility and cryopreservation was encouraging, and assays for cell functionality are underway. Figure 1. (A) The FerroBioTM hardware, shown with a cartridge in which cell isolation occurs. (B) The proportion of cells that successfully bound magnetic beads as a function of bead-cell incubation time. Final cell (C) purity and (D) viability, shown at different phases of chronological process development. Statistics: GraphPad Prism 9, 1-way ANOVA followed by a Tukey’s HSD. * p < 0.05, **** p < 0.0001. Each data point corresponds to one CBU.

Latitudinal and elevational variation in the reproductive biology of house wrens, Troglodytes aedon.

While cross-species comparisons of birds suggest that as latitude decreases or elevation increases, clutch size decreases and the duration of developmental stages and parental attentiveness increases, studies comparing populations of the same species are rare. We studied populations of house wrens, Troglodytes aedon, at high and low elevations in California and Costa Rica, collecting data on clutch size, the duration of incubation and nestling periods, parental attentiveness, nestling growth rate, and nesting success. Our data support results from cross-species comparisons, but also revealed unanticipated results from low elevation temperate zone house wrens in the southwest. This population had prolonged incubation and nestling periods similar to those found in the tropics. We also found that temperate zone females, especially those at our higher elevation site, spent more of their day incubating than did tropical females. Nest temperature at our high elevation temperate zone site was higher than that at all other tropical sites. Age at fledging did not differ between sites. Total feeding rates per chick and male feedings per chick did not vary between sites. Nest success rates showed the predicted effect of latitude, but not the predicted effects of elevation. Our results extend low elevation house wren research into the southwestern US and contribute the first intraspecific elevational comparison in the Neotropics. Data from our low elevation southwestern site present a unique suite of life history traits that align more with tropical house wrens, although with a larger clutch size, and point to food limitation and/or high predation pressure as being possible drivers of some of these differences. These results highlight the need for additional studies of house wrens and other broadly distributed species at a more diverse array of sites to better understand which forces drive the evolution of different life history strategies across major biogeographical gradients.

Direct mosquito feedings on dengue-2 virus-infected people reveal dynamics of human infectiousness.

Dengue virus (DENV) transmission from humans to mosquitoes is a poorly documented, but critical component of DENV epidemiology. Magnitude of viremia is the primary determinant of successful human-to-mosquito DENV transmission. People with the same level of viremia, however, can vary in their infectiousness to mosquitoes as a function of other factors that remain to be elucidated. Here, we report on a field-based study in the city of Iquitos, Peru, where we conducted direct mosquito feedings on people naturally infected with DENV and that experienced mild illness. We also enrolled people naturally infected with Zika virus (ZIKV) after the introduction of ZIKV in Iquitos during the study period. Of the 54 study participants involved in direct mosquito feedings, 43 were infected with DENV-2, two with DENV-3, and nine with ZIKV. Our analysis excluded participants whose viremia was detectable at enrollment but undetectable at the time of mosquito feeding, which was the case for all participants with DENV-3 and ZIKV infections. We analyzed the probability of onward transmission during 50 feeding events involving 27 participants infected with DENV-2 based on the presence of infectious virus in mosquito saliva 7-16 days post blood meal. Transmission probability was positively associated with the level of viremia and duration of extrinsic incubation in the mosquito. In addition, transmission probability was influenced by the day of illness in a non-monotonic fashion; i.e., transmission probability increased until 2 days after symptom onset and decreased thereafter. We conclude that mildly ill DENV-infected humans with similar levels of viremia during the first two days after symptom onset will be most infectious to mosquitoes on the second day of their illness. Quantifying variation within and between people in their contribution to DENV transmission is essential to better understand the biological determinants of human infectiousness, parametrize epidemiological models, and improve disease surveillance and prevention strategies.

Review: Gene of 16S rRNA is the gold standard to diagnose bacteria

The success of the biochemical tests depends on the purity of the culture, and this requires special preparations for obtaining, transporting and purifying of sample, use of selective media, duration of incubation, temperatures and others and all this takes time and effort.It may require sending slow-growing pathogens to laboratories that provide the requirements for their growth, but they may be far from the source of their collection, and thus it takes a long time to diagnose pathogen. Also, biochemical tests reflect the activity of the gene responsible for the production of a particular substance, and this gene may be present, but it does not work, and therefore the result may be a misdiagnosis. Gene of 16S rRNA is found in all bacteria, and it contains stable regions through which bacteria can be identified, and other variable regions through which bacterial species can be distinguished. As well as, diagnosis of 16S rRNA gene does not require the requirements that are required by biochemical tests for the diagnosis of bacteria, and mutations that occur in this gene do not affect its basic function, which is diagnosis, so 16S rRNA gene is the gold standard for bacterial diagnosis. In principle, it is possible to rely on phenotypic characteristics to diagnose bacteria, but confirmatory diagnosis and registration of new species is done through 16S rRNA gene using polymerase chain reaction.

Evaluation of Precision and Sensitivity of Back Extrusion Test for Measuring Textural Qualities of Cooked Germinated Brown Rice in Production Process.

The textural qualities of cooked rice may be understood as a dominant property and indicator of eating quality. In this study, we evaluated the precision and sensitivity of a back extrusion (BE) test for the texture of cooked germinated brown rice (GBR) in a production process. BE testing of the textural properties of cooked GBR rice showed a high precision of measurement in hardness, toughness and stickiness tests which indicated by the repeatability and reproductivity test but the sensitivity indicated by coefficient of variation of the texture properties. The findings of our study of the effects on cooked GBR texture of different soaking and incubation durations in the production of Khao Dawk Mali 105 (KDML 105) GBR, as measured by BE testing, confirmed that our original protocol for evaluation of the precision and sensitivity of this texture measurement method. The coefficients of determination (R2) of hardness, toughness and stickiness tests and the incubation time at after 48 hours of soaking were 0.82, 0.81 and 0.64, respectively. The repeatability and reproducibility of reliable measurements, which have a low standard deviation of the greatest difference between replicates, are considered to indicate high precision. A high coefficient of variation where relatively wide variations in the absolute value of the property can be detected indicates high sensitivity when small resolutions can be detected, and vice versa. The sensitivity of the BE tests for stickiness, toughness and hardness all ranked higher, in that order, than the sensitivity of the method for adhesiveness, which ranked lowest. The coefficients of variation of these texture parameters were 31.26, 20.59, 19.41 and 18.72, respectively. However, the correlation coefficients among the texture properties obtained by BE testing were not related to the precision or sensitivity of the test. By obtaining these results, we verified that our original protocol for the determination of the precision and sensitivity of food texture measurements which was successfully used for GBR texture measurement.

Optimization of a rapid and sensitive nucleic acid lateral flow biosensor for hepatitis B virus detection.

The utilization of direct amplification of nucleic acid from lysate has attracted interest in the advancement of straightforward and economical point-of-care assays. Consequently, this study primarily focuses on the development of a rapid, precise, and cost-effective lateral flow biosensor for the convenient detection of HBV nucleic acid at the point-of-care. Furthermore, the study evaluates the effectiveness of the direct amplification method in comparison to purified nucleic acid samples within the context of LAMP-LF biosensing approaches. The experiments conducted in this study utilized clinical serum samples that were confirmed as HBV-positive through real-time PCR assays. Sample preparation involved employing spin column nucleic acid purification and serum heat treatment. To amplify a 250bp fragment of the HBV polymerase gene, three pairs of specific LAMP primers were utilized, which were biotin-labeled and FITC-labeled for detection purposes. Various incubation temperatures (ranging from 64 to 68°C) and durations (30min, 45min, and 1h) were investigated to determine the optimal conditions for the LAMP assay. The results were subsequently assessed through fluorometric analysis, white turbidity measurements, and lateral flow assay. Milenia HybriDetect1 strips, designed for immediate use, were employed to visualize the LAMP amplicons. Furthermore, the performance of the lateral flow biosensor was evaluated using 10-fold serial dilutions of a secondary standard containing a viral load of 108 IU/ml. The optimization of the LAMP reaction was achieved at a temperature of 67°C, resulting in significant turbidity after a 30-minute incubation period. When the spin column purification method was employed, varying test bands were observed for templates ranging from 108 IU/ml to 101 IU/ml viral load. However, when serum samples underwent heat treatment and the resulting supernatant was directly used for LAMP, the lateral flow assay was capable of detecting a minimum viral load of 103 IU/ml. In resource-limited settings, the LAMP-LF assay presents a promising solution for HBV testing. However, it is important to note that direct amplification without DNA purification may diminish the performance of the approach.

Germanium (IV) Complexes with Gluconic Acid as Effectors of Penicillium tardum and Eupenicillium erubescens α-L-Rhamnosidases

α-L-Rhamnosidase (α-L-rhamnoside-rhamnohydrolase EC 3.2.1.40) showing specificity for terminal α-1,2-, α-1,4- and α-1,6-linked rhamnose residues, which often present in glycoconjugates and synthetic glycosides, can be successfully used in biotechnology for the hydrolysis of rhamnopyranoside residues present in some bioflavonoids, glycoproteins, glycolipids, and other glycoconjugates. Previously, we have shown that a significant part of the coordination compounds of various metals act as effectors of the activity of α-L-rhamnosidases. The aim of this investigation was to study the effect of a number of newly synthesized coordination compounds of Ge(IV) and Ba(II), (Co(II), Ni(II), Cu(II), Zn(II) with gluconic acid on the activity of Penicillium tardum and Eupenicillium erubescens α-L-rhamnosidases. Methods. The objects of the study were Penicillium tardum and Eupenicillium erubescens α-L-rhamnosidases. α-L-Rhamnosidase activity was determined by the Davis method using naringin as a substrate. Coordination compounds Ge(IV) and Ba(II), Co(II), Ni(II), Cu(II) ,and Zn(II) with gluconic acid were used as enzyme activity modifiers. The synthesized complexes correspond to the formulas [М(H2O)6][Ge2(OH)2(C6H8O7)2]·nH2O (М = Ba(1), n=2; Co(2), n=4; Ni(3), n=4; Cu(4), n=4; Zn(5), n=3). Results. The effect of coordination compounds 1-(5) on the activity of α-L-rhamnosidase in two strains of Penicillium tardum and Eupenicillium erubescens was studied depending on the exposure time and concentration of the effector. It was shown that compound (3) at a concentration of 0.01% (1 h incubation) led to a slight (by 5%) increase in the activity of P. tardum α-L-rhamnosidase. Compound 1 at a concentration of 0.1% led to a decrease in the activity of P. tardum α-L-rhamnosidase by 29% during the first hour, and after 24 h of incubation, a decrease in the inhibitory effect to 15% was noted. Compounds 2 and (4) activated the enzyme by 9-39% at 1h exposure. At a concentration of 0.1% and exposure time of 1 h, compound 1 increased the activity of E. erubescens α-L-rhamnosidase by 80%, while at a decrease in concentration to 0.01%, the activity increased only by 29%. In general, it should be noted that in most cases, an increase in the duration of incubation up to 24 h led to a decrease in the level of activation (or inhibition) and a return to the control values of enzyme activity. Conclusions. The variety of effects of metal coordination compounds on the activity of enzymes, depending on the nature of the cation and the origin of the enzyme, has been established. The involvement of Ba(II) had the greatest activating effect on the activity of E. erubescens α-L-rhamnosidase compared to other metals.

Biochemical Characterization of Microbial Isolates and Effect of Concentration and Incubation Time of Plant and Cow Milk Yoghurt using Response Surface Methodology

The objective of the research was to evaluate the biochemical characteristics of the various microbial isolates used in the production of plant based yoghurt, the optimal growth temperatures and the effect of incubation time and inoculum concentration on the taste and desirability of the yoghurt. Pure cultures of five (5) microbial strains (Lactobacillus acidophilus; Lactobacillus bulgaricus; Lactobacillus casei; Lactobacillus lactis; Streptococcus thermophilus) were obtained from commercial yoghurt and evaluated for their biochemical characteristics, bacterial growth temperatures and synersis. The isolated organisms were used to ferment plant-based milk product produced from blends of tigernut, Dates and Coconut. Response Surface Methodology (Face centered composite design) was used to obtain an optimized ratio formulation of 0.167 for coconut, 0.667 fortigernut and 0.167 for dates. Bacterial growth temperatures were evaluated at 4°C, 27°C, 37°C,40°C, 45°C and 60°C respectively. Results showed that full bacterial growth was noticed at temperatures of 40-45°C. The taste of the yoghurt improved from 6.58 to 6.71 when the incubation temperature changed from about 45°C to 44°C and the duration of incubation changed from 15.4 hrs to 14.7 hrs.

Characterization and in-vitro Alzheimer’s properties of exopolysaccharide from Bacillus maritimus MSM1

Four bacterial isolates were obtained from marine sediments collected from Sahl Hashish, Hurghada Red Sea, Egypt. This study was designed to search for promising anti-Alzheimer natural polysaccharide; therefore, four isolates were screened for exopolysaccharides (EPSs) production and acetylcholinesterase inhibition. The isolate S16 provided the highest EPS yield (7.51 g/L) and acetylcholinesterase inhibition. It was identified morphologically and genetically using 16S rRNA gene sequence analysis as Bacillus maritimus. A Physicochemical analysis of S16 exopolysaccharide (BMEPS) was estimated, which pointed to the presence of uronic acid and sulfate (24.7% and 18.3%, respectively). HPLC analysis indicated that mannuronic acid, glucuronic acid, glucose, and mannose are presented in a molar ratio of 0.8:1.0:2.8:2.3, respectively. Furthermore, FT-IR revealed an abundance of β-configurations. The GPC estimated the average molecular weight (Mw) as 4.31 × 104 g/mol. BMEPS inhibited AChE (IC50; 691.77 ± 8.65 μg/ ml), BChE (IC50; 288.27 ± 10.50 μg/ ml), and tyrosinase (IC50; 3.34 ± 0.09, 14.00 ± 0.14, and 22.96 ± 1.23 μg/ ml during incubation durations of 10, 20, and 40 min). It also demonstrated a selective anti-inflammatory action against COX-2 rather than COX-1. Moreover, BMEPS exhibited antioxidant capabilities as free radical and oxygen reactive species (ROS) scavenger, metal chelator, reductant agent, and lipid peroxidation suppressor. These activities are due to the distinct chemical composition. The findings of this study indicate that BMEPS could be considered as promising anti-disease Alzheimer's (AD) material in an in-vitro model, which qualifies it for advanced in-vivo studies in the discovery of alternative Alzheimer’s treatment.

Screening and characterization of agarolytic bacteria from different sources.

According to the results of our investigation, distinct bacterial isolates capable of breaking down agar were found in various nonmarine environments. The deficiency of reducing sugar in the control media demonstrates that the agar in the experiment is broken down by the bacteria to produce various oligosaccharides because the viscosity of the medium containing the agar was found to have been extremely high before inoculation, reducing with incubation duration and attaining a maximum after 48 hours. These isolates were subsequently used in tests along with additional investigation since they could create reducing sugar. Interestingly, the deterioration of agar appears to be mainly caused by Gram-negative bacteria. In order to study the agarase properties, the relative quantity of the enzyme secreted by the bacteria that hydrolyze the agar was used. The detection of extracellular agarase surrounding the colonies and the absence of stained halos on iodine-treated agar plates show that the agarase diffusing from the bacteria impacted the characteristics of the gel. Inconclusion, these agarsase-producing bacteria can be exploited for industrial applications. Waste agar from the plant tissue culture business can be utilized for a range of applications and this degraded agar can be explored for reliable and ecologically safe alternatives.

Naked neck gene and intermittent thermal manipulations during embryogenesis improve posthatch performance and thermotolerance in slow-growing chickens under tropical climates

Many studies have shown that thermal manipulations during the incubation (TMI) and naked neck gene (Na) positively affect heat-stressed broilers' thermotolerance, hatching process, and posthatch performance. Their combination could increase the beneficial effect on broilers reared under natural tropical climatic conditions. The aim of this study was to investigate the effects of the Na gene and TMI on hatching and posthatch performance of slow-growing broilers under tropical climates. The study included 1,200 hatching eggs from 2 different crosses: 1) females and males, both with a normal or fully feathered neck (na na group), and 2) females (with a normal neck) and males (bare neck) (Na na group), incubated in similar conditions until d 7. Thereafter, they were assigned to 3 subgroups for each cross: the control group (C) was incubated at standard incubation conditions (37.8°C, 60% RH). The TMI-1 group was subjected to TMI-1 (T=38.5°C, RH=65%, E10-18, 6 h/d) and TMI-2 group to TMI-2 (T=39.5°C, RH=65%, E7-16, 12 h/d). Between 450 and 504 h of incubation, eggs were checked for hatching events. During the posthatch phase, chicks from each incubation subgroups (Na na-C, Na na-TMI-1, Na na-TMI-2, na na-C, na na-TMI-1, na na-TMI-2) were raised for 12 wk at a tropical natural ambient temperature. Hatchability, hatching time, chick's temperature, final body weight (FBW), and feed conversion ratio (FCR) were determined. The results revealed that the Na gene reduced (P ˂ 0.05) hatchability. The control group had the highest mortality rate compared to TMI-1 and TMI-2 groups. There was an interaction between genotype and TMI on incubation duration, hatching weight, chick quality, FBW, and FCR (P ˂ 0.05). In conclusion, the Na gene influenced the effects of thermal manipulation. TMI-1 combined with Na gene improved the productive performances of broilers in a tropical climate.

Transmission of a bumblebee parasite is robust despite parasite exposure to extreme temperatures.

All organisms are exposed to fluctuating environmental conditions, such as temperature. How individuals respond to temperature affects their interactions with one another. Changes to the interaction between parasites and their hosts can have a large effect on disease dynamics. The gut parasite, Crithidia bombi, can be highly prevalent in the bumblebee, Bombus terrestris, and is an established epidemiological model. The parasite is transmitted between bumblebees via flowers, exposing it to a range of environmental temperatures prior to infection. We investigated whether incubation duration and temperature exposure, prior to infection, affects parasite infectivity. Prior to inoculation in B. terrestris, C. bombi was incubated at 10, 20, 30, 40 or 50°C for either 10 or 60 min. These times were chosen to reflect the length of time that the parasite remains infective when outside the host and the rate of floral visitation in bumblebees. Prevalence and infection intensity were measured in bees 1 week later. Incubation duration and the interaction between incubation temperature and duration affected the prevalence of C. bombi at 50°C, resulting in no infections after 60 min. Below 50°C, C. bombi prevalence was not affected by incubation temperature or duration. Extreme temperatures induced morphological changes in C. bombi cells; however, infection intensity was not affected by incubation duration or temperature. These results highlight that this parasite is robust to a wide range of temperatures. The parasite was not infective after being exposed to 50°C for 60 min, such temperatures likely exceed the flight abilities of bumblebees, and thus the potential for transmission. This study shows the importance of understanding the effects of environmental conditions on both hosts and parasites, which is needed to predict transmission under different environmental conditions.

Utilization of non-pathogenic bacteria to obtain optimum biofilm production for beneficial applications.

Depending on the bacteria embedded in the extracellular polymeric layer, biofilms can be advantageous or harmful. The isolated strains used in this investigation are already established to be beneficial biofilm-producing bacteria. In order to use them effectively in various domains, it is necessary to characterize them and understand their ideal physiological characteristics for maximumbiofilm growth. This study used genome sequence analysis to identify and characterize strains isolated from water samples in Raipur, Chhattisgarh, India. The nucleotide sequences were submitted to NCBI GenBank under the accession numbers Bacillus tequilensis (MN889418) and Pseudomonas beteli (MN889419) and the strains were further characterized using some advanced techniques (phase contrast microscopy, Raman spectroscopy, Fourier-transform infrared spectroscopy, and scanning electron microscope). For maximum biofilm formation by isolated bacterial strains, many physiochemical factors including incubation duration, temperature, pH, carbon source concentration, and nitrogen source concentration were further examined and optimized. The fact that these non-pathogenic strains were found in public water supplies is another important part of this research because there is a chance that they could change into pathogenic state in future and cause disease in humans.

Enzymatic Treatment of Enhanced Musa Peel Flour as Potential Low Digestible Starch


 
 
 
 Enhanced musa peel flour has a great potential as a low digestible starch that is beneficial for diabetic patients. Low digestible starch prevents the rapid increase of blood-glucose level which is crucial to protect from internal organ from damage. However, the black colour of the flour which is due to the phenolic compound oxidation in the cell wall has affected the sensory evaluation of consumers. In this work, instead of using conventional chemical treatment, enzymatic treatment is utilized to reduce the dark colour of the flour. Cell wall disruption by degradation of hemicellulose component by xylanase was hypothesized to accelerate the removal of the dark pigment. Colour reduction was optimized by varying xylanase concentration and incubation duration. Next, starch digestibility analysis was conducted using amylase enzyme to determine the starch digestibility fractions in the treated flour. As a result, instead of obtaining significant colour reduction of the flour, it is observed that there is a significant increment of resistant starch content (14%) in the treated enhanced musa peel flour. Meanwhile, reduction of 86.44% of rapidly digested starch content in the treated flour was also obtained. As a conclusion, the disruption of the cell wall by xylanase lead to the increase of the resistant starch content in the enhanced musa peel flour.
 
 
 

Synthesis, characterisation, and solution behaviour of Ag(I) bis(phenanthroline-oxazine) complexes and the evaluation of their biological activity against the pathogenic yeast Candida albicans

Three Ag(I) bis(phenanthroline-oxazine) complexes with varying lipophilicity were synthesised and characterised. The solution stoichiometry of 1:2 Ag(I):ligand was determined for each complex by the continuous variation Job’s plot method using NMR spectroscopy. NMR studies were also carried out to investigate the fluxional behaviour of the Ag(I) complexes in solution. The biological activity of the silver(I) complexes and the corresponding ligands towards a clinical strain of Candida albicans MEN was studied using broth microdilution assays. Testing showed the choice of media and the duration of incubation were key determinants of the inhibitory behaviour towards Candida albicans, however, the difference between freshly prepared and pre-prepared solutions was insignificant in minimal media. The activity of the metal-free ligands correlated with the length of the alkyl chain. In minimal media, the methyl ester phenanthroline-oxazine ligand was effective only at 60 μM, limiting growth to 67% of the control, while a 60 μM dose of the propyl ester analogue limited fungal growth at < 20% of the control. MIC50 and MIC80 values for the propyl and hexyl ester analogues were calculated to be 45 and 59 µM (propyl), and 18 and 45 µM (hexyl). Moreover, in a study of activity as a function of time it was observed that the hexyl ester ligand maintained its activity for longer than the methyl and propyl analogues; after 48 h a 60 μM dose held fungal growth at 24% of that of the control. Complexation to Ag(I) was much more effective in enhancing biological activity of the ligands than was increasing the ester chain length. Significantly no difference in activity between the three silver(I) complexes was observed under the experimental conditions. All three complexes were substantially more active than their parent ligands against Candida albicans and AgClO4 and the three silver(I) bis(phen-oxazine) complexes have MIC80 values of < 15 μM. The ability of the silver(I) complexes to hold fungal growth at about 20% of the control even after 48 h incubation at low dosages (15 μM) showcases their superiority over the simple silver(I) perchlorate salt, which ceased to be effective at dosages below 60 μM at the extended time point.

Quality attributes of set yogurt made from lactoperoxidase system activated cow’s milk

The study aimed to assess the quality attributes of set yoghurt made from cow's milk preserved by lactoperoxidase system (LPs). Fresh cow's milk was collected from Datto milk collection center and samples were grouped to control and LPs activated. Set yoghurt was prepared immediately for the controlled samples, but in LPs activated milk, sample was kept for 9 h at 25 °C ± 2 before yoghurt was made. LP enzyme was activated within 2 h after milking by adding 1 mg/ml of sodium thiocyanate (SCN_) and hydrogen peroxide (H2O2). Totally, 36 L of cow's milk was used and the experiment was done in triplicates. Titratable acidity (TA) and pH were assessed every 30 min until coagulation formed. Yoghurt samples were stored for 21 days and different parameters were assessed including incubation duration, pH, TA, microbiological quality and sensory attributes. The results revealed that LPs partially suppressed the rate of acid production during incubation of cultured milk. Yoghurt made from LPs treated milk had significantly (P < 0.05) higher pH and lower TA values. Similarly, yoghurt made from LPs treated milk had lower total bacterial counts at 24-h, 72-h and 7-days of storages. Additionally, yoghurt from LPs activated milk had lower yeast and mold counts at 24-h and 7-days. Yoghurt made from LPs activated milk received better scores for the sensory attributes. In general, LPs activated milk can be used for making set yoghurt without a negative effect. Further studies are needed to illustrate the LPs activated milk fitness in making other cultured milk products.

P-216 Evaluation of embryo quality and morphokinetics following short exposure of oocytes to spermatozoa

Abstract Study question Does embryo quality and morphokinetics following short exposure of oocytes to spermatozoa differ from conventional long exposure? Summary answer No statistical differences were found in embryo quality and morphokinetics following short exposure. Short exposure technique enables better monitoring of the embryos and easier handling. What is known already Overnight exposure of oocytes to spermatozoa may lead to toxic effects associated with reactive oxygen species. In short exposure, the duration of oocyte incubation with the sperm is reduced to 2-4 hours. Few studies showed that embryo morphology as well as pregnancy rates improved by short exposure, however, this remains controversial. Embryo morphokinetics in a time-lapse incubator yielding more accurate information on embryo quality, has not been reported for embryos derived by the short exposure technique. Study design, size, duration A retrospective study (2013-2019) 536 IVF cycles were included: 110 short exposure and 426 conventional IVF (long exposure). All embryos were cultured in the EmbryoScope. Short and long exposure cycles were compared for the average embryo scores of general and adapted laboratory in-house models. Groups were analyzed according to 2, 3 and 5 days of culture and maternal age. Moreover, differences of KID (Known implanted data) embryos morphokinetics were compared for cell division timings. Participants/materials, setting, methods IVF patients with mechanical factor, normal sperm counts. In conventional IVF, the oocytes were inseminated 6-7 hours post retrieval and mechanically denuded after 18-20 hours of incubation. In short exposure, the oocytes were inseminated and denuded 2-3 hours post retrieval and immediately placed in the EmbryoScope, for accurate annotation. In both methods oocytes were washed, incubated in EmbryoScope and cultured till the day of transfer. Women were divided by age groups: &amp;lt;35, 35-40 and &amp;gt;40. Main results and the role of chance No statistical difference was found in embryo cleavage rates and number of transferred embryos per cycle between long and short exposure in all three maternal age groups. The clinical pregnancy rate was slightly higher in the short exposure compared to the long exposure group but did not reach statistical significance (37.5% vs 32.5, p = NS). In women younger than 35y, elective single embryo transfer resulted in a statistically significant higher clinical pregnancy rate in the short exposure group compared to the long (p = 0.037). No significant differences were found in the average embryo morphokinetic score according to the general model and the adapted in-house model between the two groups. Moreover, no difference was found in embryo morphokinetics between short and long exposure in 272 KID negative and 116 KID positive embryos. Limitations, reasons for caution Since the study was retrospective and the short exposure group was smaller than the conventional IVF group, additional studies are needed to support our results. Wider implications of the findings Short exposure is more convenient and easier to perform. It is as good as long exposure in terms of clinical pregnancy rates and may be beneficial in patients younger than 35 years old. Trial registration number 0015-20-CMC