Comparative Efficacy of PercollTM Discontinuous Density Gradient Centrifugation and Glass Wool Filtration Techniques for Spermatozoa Selection in Buffalo (Bubalus bubalis)

Dead and damaged spermatozoa cells present universally in the ejaculates of all eutherian mammals exert toxic effect on contemporary healthy cells mostly through generation of excessive free radicals. This is much more evident during extended period of processing, resulting in poor ejaculate quality. The solution lies in depletion of dead/damaged spermatozoa from the neat ejaculates itself. Thus the objective of the study was to evaluate the efficiency of the protocols such as discontinuous PercollTM density gradient centrifugation (PDGC) and glass wool filtration (GWF) for depletion of dead/damaged spermatozoa from fresh semen in buffalo. Random ejaculates (n=6) of Murrah buffalo bulls were divided into two aliquots after quality assessment: PDGC and GWF protocols (Group I and II, respectively). At the end of the purification protocol, efficiency of the protocols in depleting dead/damaged spermatozoa as reflected by certain quality parameters were evaluated. The mean efficiency (%) of purification protocols based on recovery of spermatozoa was 44.68 and 40.02% for PDGC and GWF, respectively. Moreover significantly (p<0.05) greater values for quality parameters was observed in the Group II (26.4+6.8 vs 68.8+4.4 for acrosome integrity (%); 12.68+6.6 vs 57.7+7.5 for functional plasma membrane integrity (%); 20.3+5.8 vs 80.75+6.7 for viability (%) in Group I and II, respectively). It was concluded that GWF is a better technique than PGDC to filter out dead/damaged spermatozoa from fresh semen with improvement in semen quality and can be a valuable tool in assisted reproductive technology

To Study the Effect of Fertilizer and Organic Manure on Dynamic Changes of Potassium Under Cropping Sequence.

A field experiment was conducted in a farmer’s field in the Red and Laterite zone of West Bengal to evaluate the effect of integrated use of FYM and concentrated organic manures and chemical fertiliser on dynamics of potassium in soil in relation to growth and yield of component crops viz., Potato, Sesame and Rice. Three levels of chemical fertilisers viz., (i) 75% of the RDF, (ii) 100% of the RDF, and (iii) 150% of the RDF and three levels of organic manures (i) Control- applying no 0rganic manure; (ii) FYM at 20 ton/ha, and (iii) a consortium of concentrated organic manure were applied in potato after which sesame and rice were grown with the residual nutrients. Under all the three crops, application of higher doses of chemical fertilizer resulted in greater decrease in soil pH and increase in soil EC, soil OC, exchangeable, water soluble and non-exchangeable K contents of the soil. Among the two organic manures applied, the values of these parameters of the soil under all the three component crops were the highest under application of FYM. The highest pooled mean value of soil pH and lowest EC, OC, exchangeable, water soluble and non-exchangeable K contents of the soil, irrespective of the component crop in the cropping sequence, was observed under RDF75 without any organic manure and the highest under RDF150 with application of FYM. Pearson correlation co-efficient among different soil and plant parameters suggested strong relationship among themselves and K content of rice shoots, and straw of the component crops. Under limited availability of FYM, commercially available sea weed extract based concentrated organic manures could be handy and effective.

Different Methods for DNA Extraction from Yeast-Candida famata Isolated from Toddy.

Aims: Isolation and biochemical characterization of yeasts from toddy and standardization of best method for DNA extraction from yeast. Study Design: Biochemical characterization of yeast and genomic DNA extraction by manual and kits methods. Place and Duration of Study: Department of Microbiology, Institute of Genetic Engineering, Badu, kol-128, India and Molecular Mycopathology Lab, P. G. Department of Botany, Ramakrishna Mission Vivekananda Centenary College, Rahara, kol-118, India, from November 2012 –April,2013. Methodology: Toddy was collected in sterilized polythene bags from palm tree (Borassus flabellifer L; Family: Arecacea) in the morning, from Badu, 24-parganas (N) India. Isolation of yeasts was done by the method of Beech and Davenport [15] using MA (Malt extract) medium. Biochemical Identification was performed by using basal medium and procedure [1,2,15]. Genomic DNA extraction was done by manual and kits methods (UniflexTM DNA isolation Kit). Quality of extracted DNA was checked by the absorbance ratio (A260 / A280) ranged from 1.8 to 2.0. Results: By performing morphological, microscopical and biochemical characterization the isolated yeast from toddy was identified as Candida famata consulting with the key of yeast published [1,2]. The UniflexTM DNA isolation Kit method is much more convenient way to get pure and high quality DNA than the manual methods. Conclusion: Isolated yeast from toddy was identified as Candida famata. The genomic DNA of Candida famata was extracted purely by UniflexTM DNA isolation Kit. This method was better and more convenient than manual method.

In vitro assessment of drug resistance in Plasmodium falciparum in five States of India.

Background & objectives: In vitro assays are an important tool to assess baseline sensitivity and monitor the drug response of Plasmodium falciparum over time and place and, therefore, can provide background information for the development and evaluation of drug policies. This study was aimed at determining the in vitro sensitivity of P. falciparum isolates to antimalarials. Methods: The in vitro activity of 108 P. falciparum isolates obtained from five States of India was evaluated using WHO microtest (Mark III) to chloroquine, monodesethylamodiaquine, dihydroartesunate and mefloquine. Samples were collected from the States of Orissa, Jharkhand, Karnataka, Goa and Chhattisgarh from September 2007 to August 2009. In addition, representative samples from different States of India cryopreserved and culture adapted in the Malaria Parasite Bank of National Institute of Malaria Research, New Delhi, were also evaluated. Results: The proportion of isolates resistant to chloroquine and monodesethylamodiaquine was 44.4 and 25 per cent, respectively. Of the 27 isolates resistant to monodesethylamodiaquine, 16 (59.3%) were cross-resistant to chloroquine. No isolate showed resistance to dihydroartesunate and mefloquine. Isolates from Orissa showed the highest degree of resistance to chloroquine and amodiaquine followed by Jharkhand. Forty two isolates were genotyped for pfcrt T76K chloroquine resistant mutation; mutations were seen in 38 (90.47%) isolates. Interpretation & conclusions: The Indian P. falciparum isolates showed a high degree of resistance to chloroquine followed by monodesethylamodiaquine. No resistance was recorded to mefloquine and dihydroartesunate.

Retrospective study of chikungunya outbreak in urban areas of India.

Background & objectives: A retrospective study on chikungunya outbreak in India in five States viz. Delhi, Madhya Pradesh, Orissa, Maharashtra and Kerala was conducted in 2007-2008 to know the distribution and determinants of chikungunya fever outbreak in India. Methods: On the basis of high and low incidence of chikungunya fever, two districts from each State and two wards from the selected district were taken for random selection of 1000 households from 10 districts and 5 States. Semi-structured questionnaires were administered to individuals, patients, qualified health professionals and to stakeholders for collecting information. Results: The educational background and occupation of the respondents showed variations across the study States. Only in high incidence ward of Maharashtra, water storage period for 3-6 days and emptying, drying of water containers on weekly basis was noted. The study through knowledge, attitude, belief, practice (KABP) obtained individual's perception of chikungunya fever, its prevention and control. Patients’ expenditure on treatment was mainly recorded less than Rs 500 across study States. Health facility survey obtained an overview of the capacity of local health facilities. Stakeholders’ perception regarding chikungunya fever was also noted. Interpretation & Conclusions: The study revealed differences in awareness of chikungunya, cause of the disease, vector responsible, mode of transmission, biting time and elimination of breeding of mosquitoes statistically significant among high and low incidence wards of all the States. Expenditure on treatment was independent of economically active status and loss of man-days across all the States. Education and occupation did not have any relation with emptying/drying of water containers in high incidence wards. Strengthening of surveillance, information, education and communication (IEC) activities along with case management facilities may be provided by the State health department for prevention of chikungunya outbreaks in future. Stakeholders should be more involved in outbreak management and future planning.

Field evaluation of lambda-cyhalothrin (ICON 10 CS) indoor residual spraying against Anopheles culicifacies in India.

Background & objectives: Field trials of lambda-cyhalothrin 10 CS (ICON 10 CS) in indoor residual spraying (IRS) with 25 mg a.i./m2 against Anopheles culicifacies was undertaken vs malathion IRS (25% WP–2 g a.i./ m2) in Tumkur district, Karnataka; vs deltamethrin IRS (2.5% WP–20 mg a.i./m2) in Dharmapuri district; and vs lambda-cyhalothrin (10 WP–25 mg a.i./m2) in Ramanathapuram district, Tamil Nadu, India. Methods: Spray operations in the experimental villages were done by the National Institute of Malaria Research (NIMR) and in the control villages by the respective State Health Department staff. Persistence of efficacy of insecticide sprayed in villages was assessed by contact bioassays against vector mosquitoes. Entomological indicators such as per structure density, parity rates of vector mosquitoes and sporozoite rates were measured in all the three study areas using standard procedures. Mass blood surveys and active fever case detections were carried out in experimental and control villages to study the impact of IRS on malaria transmission. Results: Persistence of effectiveness of ICON 10 CS was observed up to 2–3 months in all the three study areas. ICON 10 CS was found effective at par with or better than the insecticides used in the national programme in reducing the mosquito densities and in interrupting malaria transmission in the study villages. Vector density, parity rates and malaria cases considerably reduced in the ICON 10 CS-sprayed villages. Conclusion: Field trials at three sites have established that ICON 10 CS formulation was relatively more effective than malathion 25% WP, deltamethrin 2.5% WP and lambda-cyhalothrin 10% WP in some evaluation parameters like indoor resting mosquitoes, parity rates in vector mosquitoes and persistence of effectiveness. It can be used for IRS for malaria vector control with two rounds of spray at an interval of 3 months for curtailing the malaria transmission and an additional round is recommended in perennial malaria transmission areas.

A trial on the Effects of Life Style Interventions in High-Normal Blood Pressure.

Background: This experimental (Intervention) study was conducted with objective of evaluating the outcome of non-pharmacological approach (lifestyle intervention) through reducing the modifiable risk factors on high normal blood pressure or pre-hypertension (systolic blood pressure130- 139mm of Hg and diastolic blood pressure 85-89 mm of Hg).The study was provided risk reduction management intervention was given on subjects of high normal blood pressure through adequate physical activity, tobacco cessation, dietary advice for unhealthy to healthy diet, reduction of salt intake, reduction of over weight & stress management etc. Materials and methods: The study was conducted among 434 respondents aged 30 -50 years during the period from June 2008 to May 2009 with intervention for 6 months.Random sampling, those cases were fulfilled the inclusion and exclusion. High-normal BP were found out through BP measurement & clinical examination then sample population were selected by randomization: odds numbers in study group (intervention group) and even numbers in control group (comparision group). After obtaing informed consent data were collected by interview, clinical examination, anthropometric measurement and investigations. It was three phases, first phase were case identification & intervention for study group, second phase was follow up and third phase was outcome measurement for both groups. Cases were hold a record card with contact number. Results: After six month mean difference of blood pressure, lipid profile, weight were changed both the groups but changed was significant in the study group. Mean systolic blood pressure reduced 4.1±6.0 in the study group and 1.2±6.4 in the control group,(p< 0.001). Mean diastolic pressure reduced 3.5±4.9 in study group and 1.2±6.4 in control group (p<0.001). Mean total cholesterol, LDL cholesterol and triglycerides decreased in the study group 19.7±18.6, 10.6±14.6 and 15.4 ±21 respectivley and in control group mean of all these parameters decreased to 11.5±16.5, 5.1±12.9 and 6.7±23.2 respectively. Mean HDL cholesterol for men and women in the study group was raised to 5.6±5.9 and 4.5± 6.4 respectively (p<0.001). Mean weight reduced 1.94±1.59 in study group and 0.06±1.44 in the control group (p<0.001). Conclusion: The final outcome of the study was that mean high normal blood pressure was found to reduced due to intervention of non- pharmacological management. So, If we encourages subjects after 40 years for routine health check up in hospital setting & service for risk factor detection & management (preventive cardiology), we will reduce the risk factors of high normal blood pressure (pre-hypertension) & cardiovascular diseases to some extent. Encourages population to stay with normal blood pressure & healthy life style.

Bilateral benign non functional struma ovarii with Pseudo-Meigs' syndrome.

Bilateral presentation of benign Struma ovarii is rare and has not been reported frequently in published literature. A 70-year-old postmenopausal female presented with progressive ascites, bilateral pleural effusion and elevated CA-125 levels. The contrast-enhanced computed tomography (CECT) of the abdomen and pelvis revealed a heterogenous mass in the left adnexa. These findings were suspicious for an ovarian malignancy. After surgery the diagnosis of non functional, bilateral benign Struma ovarii was made. Struma ovarii is a specialized ovarian teratoma composed predominantly of mature thyroid tissue. It is associated with pleural effusion and ascites (Pseudo-Meigs' syndrome) in 5% of cases. The combination of struma ovarii and elevated CA-125 levels has been reported infrequently. This is a rare case of bilateral benign struma ovarii associated with Pseudo-Meigs' syndrome and elevated CA-125 levels. Surgical excision of the ovarian masses induced immediate resolution of the ascites and pleural effusion and a reduction of the serum CA-125 level.

Molecular phylogeny of silk-producing insects based on 16S ribosomal RNA and cytochrome oxidase subunit I genes.

We have examined the molecular-phylogenetic relationships between nonmulberry and mulberry silkworm species that belong to the families Saturniidae, Bombycidae and Lasiocampidae using 16S ribosomal RNA (16S rRNA) and cytochrome oxidase subunit I (coxI) gene sequences. Aligned nucleotide sequences of 16S rRNA and coxI from 14 silk-producing species were used for construction of phylogenetic trees by maximum likelihood and maximum parsimony methods. The tree topology on the basis of 16S rRNA supports monophyly for members of Saturniidae and Bombycidae. Weighted parsimony analysis weighted towards transversions relative to transitions (ts, tv4) for coxI resulted in more robust bootstrap support over unweighted parsimony and favours the 16S rRNA tree topology. Combined analysis reflected clear biogeographic pattern, and agrees with morphological and cytological data.

Effect of alcohol on neurons of iso-cortex--a histomorphometric study.

Effect of chronic intake of alcohol and its subsequent withdrawal was studied in albino mice on the layers of neurons of the iso-cortex. Neuronal density per mm2 of section in different layers of iso-cortex was counted and compared in 3 groups of animals (control, ethanol fed and withdrawal). Qualitative changes on nissl granules of neurons and myelinated fibres were also studied. Mice fed with 10% ethanol v/v ad libitum for 6 months showed loss of nissl granules and nucleolus and discontinuity of nuclear membrane. Quantitatively, significant reduction in neuronal density (P<0.001) was observed in layers II+III IV and V neurons of iso-cortex. Withdrawal of ethanol for 2 months showed continued reduction of counts of neuronal density in layers II+III and V only whereas reversal of count was found significantly (P<0.001) in layer IV of iso-cortex. Qualitatively, only few neurons showed prominent nissl granules after withdrawal of ethanol. More afferent synaptic connection in layer IV may be suggested as probable factor helping relative replenishment of neuronal count after withdrawal of alcohol.

Antimicrobial susceptibility pattern of clinical isolates of non-fermentative bacteria.

152 nonfermentative bacteria were isolated from a total number of 965 clinical samples processed routinely in the laboratory of Microbiology Department, M.K.C.G Medical College in South Orissa accounting to a prevalence rate of 15.75%. Pseudomonas spp. (both pigmented and non-pigmented strains) were isolated in maximum percentage (73.6%) followed by Acinetobacter spp. (19.7%) and Alkaligenes faecalis (4.6%). Rarely encountered species were Eikenella corrodens (1.3%) and Stenotrophomonas maltophila (0.6%). Pus from various sites was the major source (116; 76%). 81% of all isolates were sensitive to amikacin and 74% to ofloxacin. Sensitivity to cefotaxime, ciprofloxacin, tobramycin, gentamicin and netlimycin ranged from 53% to 68%. Least effective drugs were carbenicillin and ceftriaxone (48% each).

Enterobacter cloacae: A predominant pathogen in neonatal septicaemia.

A total of 120 blood samples from neonates presenting with clinical signs of septicaemia were subjected for culture using brain heart infusion agar biphasic medium (BHI BPM) and glucose broth. Bacterial agents were isolated from 48 numbers (40%) of cultures. Gram-negative bacilli were isolated in maximum percentage (88.45%) of cases whereas gram-positive bacteria (coagulase negative staphylococci and group B streptococci) in 11.6% of cultures. E.cloacae (39.5%) was maximally isolated among the pathogenic bacteria followed by K.pneumoniae (23.2%), E.coli (11.6%) and others like Acinetobacter spp. (6.9%), Citrobacter freundi (4.6%) and P.mirabillis (2.3%). All the gram-negative bacilli isolates showed 100% susceptibility to amikacin, whereas 85% of E.cloacae isolates were sensitive to the same. Thus E.cloacae was found to be a predominant moderately sensitive pathogen in neonatal septicemia.

Epidemic of Vibrio cholerae serogroup O139 in Berhampur, Orissa.

During the months of May-June 2000, 194 patients with watery diarrhoea were admitted to the Infectious Diseases Ward of the M.K.C.G. Medical College, Berhampur. Ninety four rectal swabs were collected and processed according to the standard procedures. Vibrio cholerae strains were isolated from 20 samples. Of these 20 isolates, two were found to be V. cholerae O1 EITor Ogawa strain and 18 were confirmed to be V. cholerae serotype O139. All V. cholerae O139 isolates were of a single phage type (phage type 1) and the two O1 strains were of phage type 3 and phage type 27 respectively. All 20 V. cholerae isolates were positive for CAMP test, and showed uniform resistance to furazolidone and sensitivity to co-trimoxazole, amoxycillin, norfloxacin, tetracycline and gentamycin. V. cholerae O139 serotype has not been reported earlier in south Orissa. This is probably the first report of its isolation from this area.

Chemical modifications and dissociation characteristics of tyrosine and tryptophan residues in alpha-crystallin.

A quantitative estimation of surface accessibility of aromatic residues in alpha-crystallin from goat lens has been accomplished by chemical modifications using different specific reagents having varying sizes. Results of modification of tyrosine residues with N-acetylimidazole and tetranitromethane when combined with those of ionization studies carried out with hydroxyl ions having the smallest size reveal different classes of tyrosine residues in the native protein: 78 +/- 2 residues have been found to be easily available for modification; among the rest, 94 +/- 2 residues appear to be comparatively less exposed to the reagents while 28 +/- 2 residues are found to be completely unavailable for modification in the native protein and are modified only when the protein is denatured. Modification of tryptophan residues with H2O2 also indicates different classes of these residues available for oxidation at different concentrations of the oxidant. 34 +/- 2 residues of tryptophan are found to be easily oxidized at a lower concentration of H2O2 during the first phase of the reaction. The remaining tryptophan residues appear to be less exposed to the reagent. This is also corroborated from the studies of reactivities of these residues towards another specific but bulkier reagent, 2-hydroxy-5-nitrobenzyl bromide. These surface exposed aromatic residues in alpha-crystallin may be considered to be vulnerable to in vivo oxidative modifications forming insoluble aggregates which may finally contribute to the formation of cataract.

Pseudohypophosphatasia.

Two rare cases of pseudohypophosphatasia in two siblings which so far, to best of our knowledge have not been reported.