KINETICS OF THE SWELLING OF CELLS AND TISSUES

The rate of swelling of Arbacia eggs in dilute sea water, studied by Lillie and by Lucke and McCutcheon, may be expressed by the formulæ derived for the rate of increase in volume of a solution enclosed in a collodion sac. The rate of swelling of slices of carrot in distilled water, measured by Stiles and Jørgensen, may be expressed by the equation derived previously for the swelling of similarly shaped blocks of gelatin.     

Genetic Diversity of Five Local Swedish Chicken Breeds Detected by Microsatellite Markers

This study aimed at investigating the genetic diversity, relationship and population structure of 110 local Swedish chickens derived from five breeds (Gotlandshöna, Hedemorahöna, Öländsk dvärghöna, Skånsk blommehöna, and Bohuslän- Dals svarthöna, in the rest of the paper the shorter name Svarthöna is used) using 24 microsatellite markers. In total, one hundred thirteen alleles were detected in all populations, with a mean of 4.7 alleles per locus. For the five chicken breeds, the observed and expected heterozygosity ranged from 0.225 to 0.408 and from 0.231 to 0.515, with the lowest scores for the Svarthöna and the highest scores for the Skånsk blommehöna breeds, respectively. Similarly, the average within breed molecular kinship varied from 0.496 to 0.745, showing high coancestry, with Skånsk blommehöna having the lowest and Svarthöna the highest coancestry. Furthermore, all breeds showed significant deviations from Hardy-Weinberg expectations. Across the five breeds, the global heterozygosity deficit (F_IT) was 0.545, population differentiation index (F_ST) was 0.440, and the global inbreeding of individuals within breed (F_IS) was 0.187. The phylogenetic relationships of chickens were examined using neighbor-joining trees constructed at the level of breeds and individual samples. The neighbor-joining tree constructed at breed level revealed two main clusters, with Hedemorahöna and Öländsk dvärghöna breeds in one cluster, and Gotlandshöna and Svarthöna breeds in the second cluster leaving the Skånsk blommehöna in the middle. Based on the results of the STRUCTURE analysis, the most likely number of clustering of the five breeds was at K = 4, with Hedemorahöna, Gotlandshöna and Svarthöna breeds forming their own distinct clusters, while Öländsk dvärghöna and Skånsk blommehöna breeds clustered together. Losses in the overall genetic diversity of local Swedish chickens due to breeds extinction varied from -1.46% to -6.723%. The results of the current study can be used as baseline genetic information for genetic conservation program, for instance, to control inbreeding and to implement further genetic studies in local Swedish chickens.     

THE ORIENTATION OF ANIMALS BY OPPOSED BEAMS OF LIGHT

When orientation is attained under the influence of beams of parallel light opposed at 180° the deflection θ from a path at right angles to the beams is given by tan See PDF for Equation, where I ₁ and I ₂ are the photic intensities and H is the average angle between the photoreceptive surfaces. This expression is independent of the units in which I is measured, and holds whether the primary photosensory effect is proportional to I or to log I. When photokinetic side-to-side motions of the head occur, H decreases with increasing total acting light intensity, but increases if higher total light intensity restricts the amplitude of random movements; in each case, H is very nearly proportional to log I ₁ I ₂. For beams of light at 90°, See PDF for Equation. The application of these equations to some particular instances is discussed, and it is shown why certain simpler empirical formulæ previously found by others yield fair concordance with the experimental data. The result is thus in complete accord with the tropism theory, since the equations are based simply on the assumption that when orientation is attained photic excitation is the same on the two sides.     

Evidence for Natural Horizontal Transfer of the pcpB Gene in the Evolution of Polychlorophenol-Degrading Sphingomonads

The chlorophenol degradation pathway in Sphingobium chlorophenolicum is initiated by the pcpB gene product, pentachlorophenol-4-monooxygenase. The distribution of the gene was studied in a phylogenetically diverse group of polychlorophenol-degrading bacteria isolated from contaminated groundwater in Kärkölä, Finland. All the sphingomonads isolated were shown to share pcpB gene homologs with 98.9 to 100% sequence identity. The gene product was expressed when the strains were induced by 2,3,4,6-tetrachlorophenol. A comparative analysis of the 16S rDNA and pcpB gene trees suggested that a recent horizontal transfer of the pcpB gene was involved in the evolution of the catabolic pathway in the Kärkölä sphingomonads. The full-length Kärkölä pcpB gene allele had approximately 70% identity with the three pcpB genes previously sequenced from sphingomonads. It was very closely related to the environmental clones obtained from chlorophenol-enriched soil samples (M. Beaulieu, V. Becaert, L. Deschenes, and R. Villemur, Microbiol. Ecol. 40:345-355, 2000). The gene was not present in polychlorophenol-degrading nonsphingomonads isolated from the Kärkölä source.     

A continuum mechanics model for the Fåhræus-Lindqvist effect

The decrease in apparent relative viscosity that occurs when blood is made to flow through a tube whose diameter is less than about 0.3 mm is a well-known and documented phenomenon in physiology, known as the Fåhræus-Lindqvist effect. However, since the historical work of Fåhræus and Lindqvist (Amer. J. Physiol. 96(3): pp. 562–568, 1931), the underlying physical mechanism has remained enigmatic. A widely accepted qualitative explanation was provided by Haynes (Amer. J. Physiol. 198, pp. 1193–1200, 1960) according to which blood flows in microvessels with a core-annulus structure, where the erythrocytes concentrate within a central core surrounded by a plasma layer. Although sustained by observations, this conjecture lacks a rigorous deduction from the basic principles of continuum dynamics. Moreover, relations aimed to reproduce the blood apparent relative viscosity, extensively used in micro-circulation, are all empirical and not derived from the analysis of the fluid mechanical phenomena involved. In this paper, we apply the recent results illustrated in Guadagni and Farina (Int. J. Nonlinear Mech. 126, p. 103587, 2020), with the purpose of showing that Haynes’ conjecture, slightly corrected to make it more realistic, can be proved and can be used to reach a sound explanation of the Fåhræus-Lindqvist effect based on continuum mechanics. We propose a theoretical model for the blood apparent relative viscosity which is validated by matching not only the original experimental data reported by Fåhræus and Lindqvist (Amer. J. Physiol. 96(3), pp. 562–568, 1931), but also those provided by several subsequent authors.     

Use of the EM algorithm to detect QTL affecting multiple-traits in an across half-sib family analysis

QTL detection experiments in livestock species commonly use the half-sib design. Each male is mated to a number of females, each female producing a limited number of progeny. Analysis consists of attempting to detect associations between phenotype and genotype measured on the progeny. When family sizes are limiting experimenters may wish to incorporate as much information as possible into a single analysis. However, combining information across sires is problematic because of incomplete linkage disequilibrium between the markers and the QTL in the population. This study describes formulæ for obtaining MLEs via the expectation maximization (EM) algorithm for use in a multiple-trait, multiple-family analysis. A model specifying a QTL with only two alleles, and a common within sire error variance is assumed. Compared to single-family analyses, power can be improved up to fourfold with multi-family analyses. The accuracy and precision of QTL location estimates are also substantially improved. With small family sizes, the multi-family, multi-trait analyses reduce substantially, but not totally remove, biases in QTL effect estimates. In situations where multiple QTL alleles are segregating the multi-family analysis will average out the effects of the different QTL alleles.     

HYDRATION OF GELATIN IN SOLUTION

1. It was shown that the high viscosity of gelatin solutions as well as the character of the osmotic pressure-concentration curves indicates that gelatin is hydrated even at temperatures as high as 50°C. 2. The degree of hydration of gelatin was determined by means of viscosity measurements through the application of the formula See PDF for Equation. 3. When the concentration of gelatin was corrected for the volume of water of hydration as obtained from the viscosity measurements, the relation between the osmotic pressure of various concentrations of gelatin and the corrected concentrations became linear, thus making it possible to determine the apparent molecular weight of gelatin through the application of van''t Hoff''s law. The molecular weight of gelatin at 35°C. proved to be 61,500. 4. A study was made of the mechanism of hydration of gelatin and it was shown that the experimental data agree with the theory that the hydration of gelatin is a pure osmotic pressure phenomenon brought about by the presence in gelatin of a number of insoluble micellæ containing a definite amount of a soluble ingredient of gelatin. As long as there is a difference in the osmotic pressure between the inside of the micellæ and the outside gelatin solution the micellæ swell until an equilibrium is established at which the osmotic pressure inside of the micellæ is balanced by the total osmotic pressure of the gelatin solution and by the elasticity pressure of the micellæ. 5. On addition of HCl to isoelectric gelatin the total activity of ions inside of the micellæ is greater than in the outside solution due to a greater concentration of protein in the micellæ. This brings about a further swelling of the micellæ until a Donnan equilibrium is established in the ion distribution accompanied by an equilibrium in the osmotic pressure. Through the application of the theory developed here it was possible actually to calculate the osmotic pressure difference between the inside of the micellæ and the outside solution which was brought about by the difference in the ion distribution. 6. According to the same theory the effect of pH on viscosity of gelatin should diminish with increase in concentration of gelatin, since the difference in the concentration of the protein inside and outside of the micellæ also decreases. This was confirmed experimentally. At concentrations above 8 gm. per 100 gm. of H₂O there is very little difference in the viscosity of gelatin of various pH as compared with that of isoelectric gelatin.     

EFFECTS OF CYANIDE ON THE PROTOPLASM OF AMEBA

The experiments seem to indicate that the toxicity of HCN and KCN for amebæ is due to their effect on the cell membrane and not on the internal protoplasm. Concentrated solutions (N/10–N/300) of HCN or KCN produce an initial increase in viscosity of the protoplasm of amebæ (immersed) which is followed by liquefaction and disintegration of the cell. Dilute solutions of HCN or KCN decrease the viscosity of the protoplasm of amebæ. Injections of HCN or KCN into amebæ produce a reversible decrease in viscosity of the protoplasm.     

Uber Phosphorylierung im Licht

Während bisher der Zusammenhang zwischen Dehydrierung und Phosphorylierung im Licht nur für die stöchiometrischen Chinonreactionen bewiesen worden war, ist nunmehr auch für die katalytischen Chinonreactionen, die aeroben wie die anaeroben, gezeigt worden, dass die Dehydrierung die Phosphorylierung bewirkt. Eine andere Phosphorylierung, als die Phosphorylierung durch Dehydrierung gibt es in den grünen Grana nicht. Natürlich entsteht nunmehr die Frage, welche Substanz das Substrat von Dehydrierung und Phosphorylierung ist. Da die Grana keine Dunkelatmung haben und deshalb im Dunkeln nicht phosphorylieren können, auch nicht nach Zusatz von Chinon, so muss das gesuchte Substrat im Licht entstehen; und da der Photolyt der Granareactionen eine Kohlensäureverbindung ist, so muss das gesuchte Substrat eine im Licht entstehende Kohlenstoffverbindung sein. Wegen der Gärungen und dem einzigen bisher bekannten chemischen Mechanismus der Phosphorylierung durch Dehydrierung (4) denkt man an Triosephosphat. Aber chemisch ist jeder Aldehyd möglich.     

The three-dimensional structure of alpha1-purothionin in solution: combined use of nuclear magnetic resonance, distance geometry and restrained molecular dynamics

The determination of the three-dimensional solution structure of α1-purothionin using a combination of metric matrix distance geometry and restrained molecular dynamics calculations based on n.m.r. data is presented. The experimental data comprise complete sequence-specific proton resonance assignments, a set of 310 approximate interproton distance restraints derived from nuclear Overhauser effects, 27 Ø backbone torsion angle restraints derived from vicinal coupling constants, 4 distance restraints from hydrogen bonds and 12 distance restraints from disulphide bridges. The average atomic rms difference between the final nine converged structures and the mean structure obtained by averaging their coordinates is 1.5 ± 0.1 å for the backbone atoms and 2.0 ± 0.1 å for all atoms. The overall shape of α1-purothionin is that of the capital letter L, similar to that of crambin, with the longer arm comprising two approximately parallel α-helices and the shorter arm a strand and a mini anti-parallel β sheet.     

Isolation and characterization of a highly effective bacterium Bacillus cereus b-525k for hexavalent chromium detoxification

The chromate resistant Gram-positive Bacillus cereus strain b-525k was isolated from tannery effluents, demonstrating optimal propagation at 37 °C and pH 8. The minimum inhibitory concentration (MIC) test showed that B. cereus b-525k can tolerate up to 32 mM Cr⁶⁺, and also exhibit the ability to resist other toxic metal ions including Pb²⁺ (23 mM), As³⁺ (21 mM), Zn²⁺ (17 mM), Cd²⁺ (5 mM), Cu²⁺ (2 mM), and Ni²⁺ (3 mM) with the resistance order as Cr ⁶⁺ > Pb²⁺ > As³⁺ >Zn²⁺ >Cd²⁺ >Ni²⁺ >Cu²⁺. B. cereus b-525k showed maximum biosorption efficiency (q) of 51 mM Cr⁶⁺/g after 6 days. Chromate stress elicited pronounced production of antioxidant enzymes such as catalase (CAT) 191%, glutathione transferase (GST) 192%, superoxide dismutase (SOD) 161%, peroxidase (POX) 199%, and ascorbate peroxidase (APOX) (154%). Within B. cereus b-525k, the influence of Cr⁶⁺ stress (2 mM) did stimulate rise in levels of GSH (907%) and non-protein thiols (541%) was measured as compared to the control (without any Cr⁶⁺ stress) which markedly nullifies Cr⁶⁺ generated oxidative stress. The pilot scale experiments utilizing original tannery effluent showed that B. cereus b-525k could remove 99% Cr⁶⁺ in 6 days, thus, it could be a potential candidate to reclaim the chromate contaminated sites.     

LABYRINTH AND EQUILIBRIUM : I. A COMPARISON OF THE EFFECT OF REMOVAL OF THE OTOLITH ORGANS AND OF THE SEMICIRCULAR CANALS.

1. A dogfish from which all six ampullæ have been removed maintains its equilibrium; the righting reactions occur promptly; compensatory movements of the eyes occur in response to rotations in all planes except the horizontal; the compensatory position of the eyes is retained if the animal is held in an abnormal position. Both the static and dynamic functions of equilibrium continue, therefore, after complete removal of all the semicircular canals and all the ampullæ. 2. After complete removal of the otoliths from the vestibules without injury to the ampullæ the animal maintains its equilibrium in the water, rights itself promptly, and makes compensatory motions to rotations in all planes. If held in an abnormal position the compensatory position of the eyes is maintained. Both static and dynamic functions of equilibrium continue. 3. Destruction of both the semicircular canals and the otolith organs completely abolishes all compensatory movements and equilibrium reactions of labyrinthine origin. 4. It is pointed out that these observations do not justify the theory of Mach and Breuer that the ampullæ and semicircular canals are the organs for the dynamic functions of equilibrium, and that the otoliths are the organs for the static functions of equilibrium. 5. The new experiments recorded in this paper show that the ampullæ alone (without the otoliths) suffice for all the dynamic and all the static functions of equilibrium of the ear; and that the otolith organs (without the ampullæ) suffice for all the static and for all the dynamic functions of equilibrium of the ear with the exception of the response to a rotation of the animal in a horizontal plane.     

RELATION BETWEEN THYROID GLAND,METAMORPHOSIS, AND GROWTH

1. Two substances are involved in amphibian metamorphosis as studied in Ambystoma opacum: first, iodine, which is taken up by the food, and second, an excretor substance, which is evolved during the processes of growth and serves to induce the excretory function of the thyroid gland. 2. This explains why in larvæ, whose metamorphosis is inhibited by lack of iodine, growth is checked at the time when metamorphosis should occur; for at this time the excretor substance commences to act and this results, if iodine is absent, in the excretion by the thyroid of toxic substances which cause the breakdown of proteins and consequently a decrease in size of the larvæ. 3. Larvæ whose metamorphosis is inhibited by extirpation of the thyroid or by the hereditary lack of a thyroid (as is the case in Typhlomolge) can grow normally, since in them the action of the excretor substance cannot result in the excretion by the thyroid of a toxic growth-inhibiting substance. 4. At low temperature less excretor substance is produced than at high temperature during an equal rate of growth; therefore larvæ kept at low temperature reach a larger size than larvæ kept at high temperature, before they metamorphose.     

Review of genus Pseudorthocladius Goetghebuer, 1943 (Diptera,Chironomidae) from China

The genus Pseudorthocladius Goetghebuer, 1943 from China, including 12 species, is reviewed. Five new species, P. (P.) binarius sp. n., P. (P.) cylindratus sp. n., P. (P.) digitus sp. n., P. (P.) ovatus sp. n. and P. (P.) paucus sp. n. are described and illustrated as adult males. P. (P.) cristagus Stur & Sæther, 2004, P. (P.) jintutridecima (Sasa, 1996), P. (P.) macrovirgatus Sæther & Sublette, 1983, P. (P.) morsei Sæther & Sublette, 1983, P. (P.) uniserratus Sæther & Sublette, 1983, P. (L.) wingoi Sæther & Sublette, 1983 are newly recorded in Oriental Region. A key to the males of Pseudorthocladius in China is presented.     

Bioinformatics with a French accent

A report on the bioinformatics conference 'JOBIM: Journées ouvertes biologie informatiques mathématiques', Lyon, France, 6-8 July 2005.     

Inactivation of Enteric Adenovirus and Feline Calicivirus by Chlorine Dioxide

Chlorine dioxide (ClO₂) inactivation experiments were conducted with adenovirus type 40 (AD40) and feline calicivirus (FCV). Experiments were carried out in buffered, disinfectant demand-free water under high- and low-pH and -temperature conditions. Ct values (the concentration of ClO₂ multiplied by contact time with the virus) were calculated directly from bench-scale experiments and from application of the efficiency factor Hom (EFH) model. AD40 Ct ranges for 4-log inactivation (Ct_99.99%) at 5°C were >0.77 to <1.53 mg/liter × min and >0.80 to <1.59 mg/liter × min for pH 6 and 8, respectively. For 15°C AD40 experiments, >0.49 to <0.74 mg/liter × min and <0.12 mg/liter × min Ct_99.99% ranges were observed for pH 6 and 8, respectively. FCV Ct_99.99% ranges for 5°C experiments were >20.20 to <30.30 mg/liter × min and >0.68 mg/liter × min for pH 6 and 8, respectively. For 15°C FCV experiments, Ct_99.99% ranges were >4.20 to <6.72 and <0.18 mg/liter × min for pH 6 and 8, respectively. Viral inactivation was higher at pH 8 than at pH 6 and at 15°C than at 5°C. Comparison of Ct values and inactivation curves demonstrated that the EFH model described bench-scale experiment data very well. Observed bench-scale Ct_99.99% ranges and EFH model Ct_99.99% values demonstrated that FCV is more resistant to ClO₂ than AD40 for the conditions studied. U.S. Environmental Protection Agency guidance manual Ct_99.99% values are higher than Ct_99.99% values calculated from bench-scale experiments and from EFH model application.     

Distantiae Transmission of Trypanosoma cruzi: A New Epidemiological Feature of Acute Chagas Disease in Brazil

Background

The new epidemiological scenario of orally transmitted Chagas disease that has emerged in Brazil, and mainly in the Amazon region, needs to be addressed with a new and systematic focus. Belém, the capital of Pará state, reports the highest number of acute Chagas disease (ACD) cases associated with the consumption of açaí juice.

Methodology/Principal Findings

The wild and domestic enzootic transmission cycles of Trypanosoma cruzi were evaluated in the two locations (Jurunas and Val-de Cães) that report the majority of the autochthonous cases of ACD in Belém city. Moreover, we evaluated the enzootic cycle on the three islands that provide most of the açaí fruit that is consumed in these localities. We employed parasitological and serological tests throughout to evaluate infectivity competence and exposure to T. cruzi. In Val-de-Cães, no wild mammal presented positive parasitological tests, and 56% seroprevalence was observed, with low serological titers. Three of 14 triatomines were found to be infected (TcI). This unexpected epidemiological picture does not explain the high number of autochthonous ACD cases. In Jurunas, the cases of ACD could not be autochthonous because of the absence of any enzootic cycle of T. cruzi. In contrast, in the 3 island areas from which the açaí fruit originates, 66.7% of wild mammals and two dogs displayed positive hemocultures, and 15.6% of triatomines were found to be infected by T. cruzi. Genotyping by mini-exon gene and PCR-RFLP (1f8/Akw21I) targeting revealed that the mammals and triatomines from the islands harbored TcI and Trypanosoma rangeli in single and mixed infections.

Conclusion/Significance

These findings show that cases of Chagas disease in the urban area of Belém may be derived from infected triatomines coming together with the açaí fruits from distant islands. We term this new epidemiological feature of Chagas disease as “Distantiae transmission”.     

Arthrite rhumatoide traitée par noréthynodrel associée à mestranol: Aspects cliniques et tests de laboratoire

A cause de l''effet favorable de la grossesse sur l''activité de l''arthrite rhumatoïde, on s''est demandé si une pseudo-grossesse, produite par un progestatif de synthèse n''entraînerait pas une rémission du moins partielle de cette maladie.Noréthynodrel associée à mestranol (Enovid), 30 mg./jour, a été administrée à 44 femmes pendant quatre à 24 mois. A cause d''effets secondaires indésirables, 11 patientes furent soustraites de l''investigation. Les résultats s''appuient sur 33 cas. Une rémission apparente complète s''est manifestée chez sept patientes aux stades précoces de la maladie; chez 15, une amélioration objective des signes inflammatoires a été observée; chez quatre, une amélioration subjective seule a été notée; chez sept, il n''y a eu aucune amélioration. Dix-sept femmes sur 36 ont présenté une altération d''un ou plusieurs tests hépatiques. Trois présentèrent un ictère cholestatique. Les 17-OH plasmatiques se sont élevés à trois ou quatre fois la normale.De cette étude il ressort que noréthynodrel associée à mestranol peut produire une atténuation des signes inflammatoires de l''arthrite rhumatoïde. L''effet est palliatif, mitigé et non curatif et ne résulte pas nécessairement de l''état de pseudo-grossesse en soi.     

Sensitivity of Plasmodium falciparum to Antimalarial Drugs in Hainan Island,China

Pyronaridine and artesunate have been shown to be effective in falciparum malaria treatment. However, pyronaridine is rarely used in Hainan Island clinically, and artesunate is not widely used as a therapeutic agent. Instead, conventional antimalarial drugs, chloroquine and piperaquine, are used, explaining the emergence of chloroquine-resistant Plasmodium falciparum. In this article, we investigated the sensitivity of P. falciparum to antimalarial drugs used in Hainan Island for rational drug therapy. We performed in vivo (28 days) and in vitro tests to determine the sensitivity of P. falciparum to antimalarial drugs. Total 46 patients with falciparum malaria were treated with dihydroartemisinin/piperaquine phosphate (DUO-COTECXIN) and followed up for 28 day. The cure rate was 97.8%. The mean fever clearance time (22.5±10.6 hr) and the mean parasite clearance time (27.3±12.2 hr) showed no statistical significance with different genders, ages, temperatures, or parasite density (P>0.05). The resistance rates of chloroquine, piperaquine, pyronarididine, and artesunate detected in vitro were 71.9%, 40.6%, 12.5%, and 0%, respectively (P<0.0001). The resistance intensities decreased as follows: chloroquine>piperaquine>pyronarididine>artesunate. The inhibitory dose 50 (IC₅₀) was 3.77×10^-6 mol/L, 2.09×10^-6 mol/L, 0.09×10^-6 mol/L, and 0.05×10^-6 mol/L, and the mean concentrations for complete inhibition (CIMC) of schizont formation were 5.60×10^-6 mol/L, 9.26×10^-6 mol/L, 0.55×10^-6 mol/L, and 0.07×10^-6 mol/L, respectively. Dihydroartemisinin showed a strong therapeutic effect against falciparum malaria with a low toxicity.     

DNA barcoding and genetic fidelity assessment of micropropagated Aenhenrya rotundifolia (Blatt.) C.S. Kumar and F.N. Rasm.: a critically endangered jewel orchid

Aenhenrya rotundifolia is a critically endangered terrestrial jewel orchid. It is monotypic and endemic to evergreen forests of southern western ghats of India. In the present study, identification of this plant species is validated with DNA barcoding using matK and rbcL chloroplast markers. Further, germ-free juvenile axillary bud explants were cultured on Mitra medium supplemented with different kinds of cytokinins like 6-benzyladenine, 6-furfurylaminopurine, N⁶-(Δ²-isopentyl) adenine, thidiazuron, zeatin and meta-topolin as well as auxins such as α-naphthaleneacetic acid, indole-3-acetic acid and indole-3-butyric acid at different concentrations and combinations for successful proliferation and establishment in vitro. After 12 weeks of culture, axillary bud explants produced an average of 30.12 ± 0.71 shoots per explant, 3.87 ± 0.06 cm shoot length, 1671 ± 2.82 mg fresh mass of proliferated shoots with a proliferation frequency of 100% on Mitra medium supplemented with 6.20 µM meta-topolin and 2.25 µM thidiazuron. No root formation was observed in in vitro proliferated microshoots. However, tiny hair like projections were observed in some elongated shoots on Mitra medium pertaining to 5.37 µM NAA. The tiny hair like structure bearing plantlets were hardened and acclimatized with 100% survival rate in the polytunnel chamber. After 8–10 months of establishment ex vitro, flowering was observed. Additionally, the genetic fidelity of in vitro derived plants was tested with ISSR and SCoT marker profiling. The test results revealed that the plants derived from the protocol has 99% genetic similarity to that of the donor mother plant. This study can be applied in forensic interventions of this species, describes the maintenance of germplasm in vitro and establishment of new viable population in its original habitats by restoring existing sites of this critically endangered jewel orchid.