Validation of the Use of Dried Blood Samples for the Detection of Toxoplasma gondii Antibodies in Stray Cats (Felis s. catus)

If validated beforehand, the analysis of dried blood on blotting paper (BP samples) is very useful for monitoring free-ranging animals. We aimed to validate this method for the detection of antibodies against Toxoplasma gondii in stray cats. We used the modified agglutination test (MAT) in 199 sample pairs of sera and BP samples from 54, 39, 56, and 50 cats trapped during four periods in five dairy farms. Screening was at 1:6, 1:12, and 1:24 dilutions. The cut-off value was at MAT titre ≥ 24, but MAT titre ≥ 12 was also considered for BP samples that often have a higher dilution level. Depending on the period, sample type, and cut-off value, sensitivity of the analysis of the BP sample vs. serum varied from 87.1% to 100% and specificity ranged from 72.22% to 100%. The concordance values and Kappa coefficient showed a substantial to excellent agreement between the results of the two methods, whatever the cut-off value. These findings quantifiably validate the use of MAT on BP samples for the detection of antibodies to T. gondii in stray cats, but we recommend expressing results from BP samples with several cut-off values as the MAT titres tend to be lower than those of sera.

Bone Preparation from Embalmed Human Cadavers - A Retrieval and Curation Technique

Bone preparation involves soft tissue removal, maceration, bleaching and labelling. In the absence of a standardized methodology a large repository of human bones are lost, as most medical colleges do not process bones after the dissection of human cadavers. The present study therefore conducted with the aim of evaluating the least time-consuming and effective method of bone preparation from embalmed and wet specimens. The method used included a process of maceration, which involves soft tissue removal and then boiling the bones in 60 litres of water for 2 hours. The process of maceration was augmented by adding potassium hydroxide pellets (caustic potash mol. wt. 56.11) after 30 minutes of initiation of boiling; 200-250 gm in the case of male bones and 150 – 200 gm in the case of female bones. After maceration was complete, the bones were bleached by soaking them in 30 – 35 litres of hydrogen peroxide 30% w/v solution (mol. wt. 34.01) for 12-14 hours. The bleached bones were then washed with water and soaked in 30 -35 litres of acetone (extra pure mol. wt. 58.08, boiling point 55.5° – 56°C) for 12 hours to degrease them. The bones dried naturally by spreading them on blotting paper and subsequently painted with a mixture of half a litre of lacquer and half a litre of lacquer thinner. This study concluded that the preparation of bones using the above method was effective, fast, odourless, and good quality human bones for anatomical study resulted.

A practical, low-cost, short-term storage method for genomic DNA

The aim of this study was to assess the DNA preservation capability of cellulose paper towel and blotting paper as low-cost alternatives to commercial DNA preservation products. Chicken blood was applied as DNA source to each paper towel, blotting paper, FTA® cards and DNA/RNA Shield™. All samples were stored at room temperature for 130 days. DNA extraction from dried blood spots was performed after various time periods using Tris–EDTA and NaOH protocols. PCR activity and the mean amount of DNA isolated from paper towels were reliable. The results of this study demonstrated that cellulose-based blotting paper and especially paper towel had considerable DNA binding and preservation capacity for at least 130 days at room temperature without DNA degradation.

Comparison of Immunofluorescence Assays Results for Diagnosis of Rickettsial Diseases Using Direct Serum and Filter Paper Serum Spots

Background: Rickettsial diseases are difficult to diagnose in Bangladesh due to low index of suspicion and lack of diagnostic facilities in most laboratories especially in rural areas. Immunofluorescence assay (IFA) is the gold standard serological test for diagnosis of Rickettsial diseases. For serological diagnosis of rickettsial diseases, use of filter paper is an inexpensive and convenient method for collecting, storing, and transporting of blood samples with little space without refrigeration. Current study shows that the rickettsial diseases were present in febrile patients in Bangladesh. The study also compared the results of direct serum by IFA and filter paper serum spot followed by IFA as an alternative way of testing frozen samples. Materials and Methods: This observational study for evaluation of rickettsial diseases from direct serum by IFA and sera dried on blotting paper followed by IFA at the department of Virology, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka, Bangladesh. A total of 152 patients’ sera were selected by systemic random sampling of every 3rd sera of 556 febrile patients who were tested negative for dengue and or Chikungunya virus infection. Forty cases were confirmed by IFA from direct sera. In addition, 60 samples were selected from 152 sera by systemic random sampling of every 3rd sera for blotting on filter paper for further IFA test. Out of 60 sera, 35 were confirmed by IFA from direct sera and of the 35 confirmed positive cases, 31(88.6%) were positive by IFA after blotting paper assay. Taking the IFA of direct sera as gold standard test, the sensitivity, specificity, positive predictive value and negative predictive value of IFA from blotting paper eluted sera were 88%, 100%, 100% and 86% respectively. Conclusion: The study showed the use of filter paper is an inexpensive and convenient method for storing and transporting samples for serological tests like IFA in diagnosis of rickettsial diseases. J Dhaka Medical College, Vol. 28, No.2, October, 2019, Page 153-158

EMPLOYMENT OF NOT STANDARDIZED CONTAINERS BY RST AND THEIR INTERFERENCE IN THE PHYSIOLOGICAL QUALITY OF SEEDS OF Pinus elliottii

The determining factor for ensuring success in a forest enterprise is directly linked to the quality of the seeds used in the nursery. In this sense, the objective of this study was to evaluate the physiological quality of Pinnus Elliotis in different containers compared to the standardized test by RST. Cellboxes (tissue culture boxes), with different diameters and cell numbers, respectively (0.7, 1.1, 16 and 21 mm and 96, 48, 24, 12) with circular blotting paper substrates were used. The experimental design used was a completely randomized with 5 treatments and 8 repetitions. The treatment (T1) showed the highest average germination and the lowest fungal infestation, compared to the gerbox container. Cellbox containers showed better values for germination, showing that the division of cells positively influences the health quality of Pinus elliottii Engelm var. elliottii.

Germination and vigor of Encholirium spectabile seeds according to geographical region, substrate and sowing position

Given the potential for commercial exploitation of E. spectabile and aiming to curb predatory extractivism, the development of researches that guide its cultivation becomes relevant. This study aims to evaluate the effects of different substrates and seed positions at sowing on the germination and vigor of seeds from two geographical regions. The experiment design was completely randomized in a 2 x 4 x 2 factorial design (seeds from two geographic regions: i.e., Serra Talhada-PE and Graça-CE, both in Brazil; four substrates: blotting paper, sand, vermiculite and coconut fiber: and two sowing positions: over and in between the substrate). Sowing on blotting paper, sand, vermiculite and coconut fiber, in general, was favorable to germination and germination speed index for seeds from both geographic regions. Sowing in between coconut fiber was detrimental to germination and root development regardless of the seed geographical region. However, sowing in between and over coconut fiber and in between sand favored shoot growth. The best combinations for dry matter production were sowing over paper and sand. The substrates blotting paper, sand, coconut fiber and vermiculite are favorable to E. spectabile seed germination regardless of geographic regions, provided that the sowing is made over the substrates.

Is a long hygroscopic awn an advantage for Themeda triandra in drier areas?

Themeda triandra has bigeniculate hygroscopic lemma seed awns that twist when wet and drying, thereby transporting the caryopsis across the soil surface into suitable germination microsites. The prediction that awns would be longer in drier grassland and have greater motility to enable them to move quickly and further to find scarce germination sites was tested in KwaZulu-Natal. Awns (n = 100) were collected from 16 sites across a mean annual precipitation gradient (575-1223 mm), ranging from 271-2097 m a.s.l. The daily movement of hydrated long and short awns (n = 10) across blotting paper was tracked for five days, and the rotational speed of anchored awns was measured. Awn length varied considerably (mean: 41.4-63.2 mm; sd: 3.44-8.99) but tended to increase (r = 0.426, p = 0.099) not decline, with increasing MAP. Awn length was unrelated to elevation, temperature and aridity indices. Long awns rotated at the same rate (2 min 48 sec) but moved twice as fast (46.3 vs. 22.1 mm day-1) and much further (maximum: 82.1 vs. 38.6 mm day-1) than short awns. Whether moisture limits awn development, the benefit of longer awns to negotiate densely tufted mesic grassland, and the multifunctionality of awns require investigation.

A new method for the direct tracking of in vivo lignin nanocapsules in Eragrostis tef (Poaceae) tissues

Environmental concerns have driven scientists to research new eco-friendly approaches for the preparation of nanosystems. For this purpose, novel bio-polymers have been selected. Among these, one of the most promising is lignin, which is biodegradable and biocompatible. Additionally, lignin is one of the main by-products of the paper industry and can be re-used in nanosystems building. Lignin-based nanosystems could be used in agriculture, to improve the uptake of bioactive compounds, thus avoiding soil pollution. However, the mechanism of penetration in the plant and the route of transportation within the internal plant tissues are unknown and need to be clearly elucidated. Here we present a method of lignin nanocapsules staining and tracking by fluorochrome: Fluoral Yellow 088, which is a well-suited dye for the tracking of lipids and other oil phases. Two different applications were applied: in the first one fourteen-day plants were soaked with fluorescent nanocapsules (fNCs) pure solution and in the second one, Eragrostis tef plants were laid down on blotting paper and soaked with diluted fNCs solution. Wetting the roots of Teff plantlets with the pure fNCs solution resulted in the most efficient way of nanocapsule entrance. The dyeing of lignin nanocapsules allowed us to track them in Eragrostis tef plant tissues through microscopic observations. In particular, fNCs were proven to be able to permeate roots, reaching xylem vessels where, through water pressure, they reached the leaf.

Characterizing the fluid–matrix affinity in an organogel from the growth dynamics of oil stains on blotting paper

Fluid–matrix affinity in an organogel is characterized by capillarity-induced oil release using absorbing paper.