Search

Diabetic retinopathy (DR) is an important microvascular complication of diabetes mellitus (DM), causing significant visual impairment worldwide. Current gold standards for retarding the progress of DR include blood sugar control and regular fundus screening. Despite these measures, the incidence and prevalence of DR and vision-threatening DR remain high. Given its slowly progressive course and long latent period, opportunities to contain or slow DR before it threatens vision must be explored. This narrative review assesses the recently described unconventional strategies to retard DR progression. These include gut-ocular flow, gene therapy, mitochondrial dysfunction-oxidative stress, stem cell therapeutics, neurodegeneration, anti-inflammatory treatments, lifestyle modification, and usage of phytochemicals. These therapies impact DR directly, while some of them also influence DM control. Most of these strategies are currently in the preclinical stage, and clinical evidence remains low. Nevertheless, our review suggests that these approaches have the potential for human use to prevent the progression of DR.

Conjunctival swabs (CS) are the major source of sampling for ocular microbiome studies, however collecting CS from the diseased eyes is difficult and painful. In this study, as an alternative to CS, a less invasive approach of tear collection was used to establish the bacterial microbiome in healthy eyes. Tear bacterial microbiome was generated from the DNA of tears (n = 24; male = 16 and female = 8) of healthy volunteers aged from 20 to 52 years. Sequencing of V3-V4 region of 16S rRNA gene was performed on the Illumina platform. Reads were processed in QIIME to assign the taxa. Statistical analysis of the tear microbiome was done in R to assess the alphadiversity and betadiversity indices. Tear microbiome was generated in all the 24 tear samples. Eight out of the top 10 predominant bacterial genera remained same in both tear and CS microbiomes, which include genera such as Corynebacterium, Staphylococcus, Streptococcus, Mycobacterium, Escherichia-Shigella, Lactobacillus, Bacillus and Acinetobacter. The similarity network analysis indicates that 144 out of 145 genera of tear cohort matched with conjunctival swabs. However, tear and CS microbiomes differed in the abundance of the predominant bacterial genera. The bacterial microbiome of tears in adults appears to be stable and is comparable with that of CS microbiome.

We investigated the link between polymorphisms in genes involved in mitochondrial biogenesis, mitochondrial transcription factor A (TFAM) and Peroxisome proliferator activated receptor gamma coactivator-1 alpha (PGC-1α) and further studied the role of these genes on the pathophysiology of polycystic ovary syndrome (PCOS). This case-control study was carried out in 118 PCOS cases and 110 controls. In the present study we genotyped three polymorphisms of PGC1-α gene (rs8192678-Gly482Ser, rs13131226 and rs2970856) and polymorphism of TFAM gene (rs1937-+35G/C) by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis. In addition, to better understand genetic contributions to the pathophysiology of PCOS, mtDNA copy number (MCN) was quantified using a qRT-PCR assay in the subjects. The results revealed that the distribution of genotypes and allele frequency of the PGC-1α Gly482Ser polymorphism in PCOS patients was statistically significant from those of the control group respectively (OR-2.488; 95% CI-1.0673 to 5.7998; P = 0.047), (OR-1.6091; 95% CI-1.0955 to 2.3634; P = 0.015) indicating that the presence of ‘A’ allele might confer risk to PCOS. Patients with the ‘AA’ genotype showed significantly lower levels of MCN compared with patients with other genotypes. In addition, patients carrying CT genotype of PGC1-α rs2970856 demonstrated significantly higher levels of LH ( P = 0.030) than TT and CC genotypes. In conclusion, our study indicates that carriers of the PGC-1α rs8192678 ‘Ser’ allele have increased risk of developing PCOS.

The aim of the study was to identify a cryo-device that would be best suited for the vitrification of buffalo immature cumulus-oocyte complexes (COCs) as judged by viability and meiotic competence of the vitrified-warmed oocytes and their development ability following in vitro fertilization (IVF). The expression of oocyte secreting factors and their receptors (GDF9, BMP15, BMPR2, TGFBR1) and apoptosis related genes (BCL2, BAX, P53, C-MYC) were compared in vitrified-warmed oocytes after in vitro maturation. COCs from the ovaries of slaughtered buffaloes were vitrified in a combination of dimethyl sulfoxide, ethylene glycol, and sucrose using either a conventional straw (CS), open pulled straw (OPS), cryoloop (CL), hemistraw (HS) or cryotop (CT). The fresh COCs were exposed to vitrification and warming solutions as in other vitrification methods without plunging in to liquid nitrogen (EC). The viability of vitrified-warmed COCs, 2 h post warming in HS and CT was similar to fresh and EC groups but significantly higher than CS and OPS methods. The proportions of oocytes with first polar body after 24 h in vitro maturation were significantly higher in HS and CT methods than in CS, OPS and CL methods. The development ability of these vitrified-warmed oocytes to blastocyst stage following IVF in all vitrified groups was significantly lower than control and EC groups. Among the vitrified groups, the blastocyst rate in HS, CT and CL groups was significantly higher than in OPS and CS groups. It was also observed that the expression levels of GDF9, BMP15, BMPR2, TGFBR1, BCL2, BAX, P53 and C-MYC genes in vitrified-warmed COCs in CT, HS and CL groups were similar to control. The results indicated that HS, CT and CL are more suitable cryo-devices for vitrification of buffalo immature oocytes.

Background The review focuses on the bacteria associated with the human eye using the dual approach of detecting cultivable bacteria and the total microbiome using next generation sequencing. The purpose of this review was to highlight the connection between antimicrobial resistance and biofilm formation in ocular bacteria. Methods Pubmed was used as the source to catalogue culturable bacteria and ocular microbiomes associated with the normal eyes and those with ocular diseases, to ascertain the emergence of anti-microbial resistance with special reference to biofilm formation. Results This review highlights the genetic strategies used by microorganisms to evade the lethal effects of anti-microbial agents by tracing the connections between candidate genes and biofilm formation. Conclusion The eye has its own microbiome which needs to be extensively studied under different physiological conditions; data on eye microbiomes of people from different ethnicities, geographical regions etc. are also needed to understand how these microbiomes affect ocular health.

Microbial keratitis is an infectious disease of the eye, in which the cornea is inflamed. Under severe conditions, keratitis can lead to significant loss of vision and enucleation of the eye. Ocular trauma is the major risk factor causing keratitis and microorganisms viz., bacteria, fungi, viruses are the causative agents. The current study characterized the conjunctival bacterial microbiomes of healthy individuals and individuals with bacterial keratitis (BK) and assessed whether ocular microbiome dysbiosis is prevalent in BK patients. Ocular bacterial microbiomes were generated from the conjunctival swabs of healthy controls (HC-SW) and conjunctival swabs (BK-SW) and corneal scrapings (BK-CR) of BK patients using V3-V4 amplicon sequencing and data analysed using QIIME and R software. The Alpha diversity indices, diversity and abundance of different phyla and genera, heat map analysis, NMDS plots and inferred functional pathway analysis clearly discriminated the bacterial microbiomes of conjunctival swabs of healthy controls from that of BK patients. Preponderance of negative interactions in the hub genera were observed in BK-CR and BK-SW compared to HC-SW. In addition, a consistent increase in the abundance of pathogenic bacteria, as inferred from published literature, was observed in the conjunctiva of BK patients compared to HC and this may be related to causing or exacerbating ocular surface inflammation. This is the first study demonstrating dysbiosis in the ocular bacterial microbiome of conjunctiva of bacterial keratitis patients compared to the eye of healthy controls. The bacterial microbiome associated with the corneal scrapings of keratitis individuals is also described for the first time.

Objective Polycystic ovarian syndrome (PCOS) is a complex and multifactorial disorder believed to be the consequence of a complex interaction between genetic, immunological, and environmental factors. The main aim of this study was to investigate the association of Connexin37 ( Cx37 )/Gap junction alpha 4 (GJA4) gene C1019T single nucleotide polymorphism (SNP) with the susceptibility to polycystic ovarian syndrome (PCOS) in South Indian women. Study design This study comprises 98 PCOS patients and 100 healthy women without PCOS of South Indian origin. We genotyped total of seventeen selected Cx37 SNPs including C1019T (rs1764391) by polymerase chain reaction and sequencing analysis. The genotype frequency and allele distributions of cases and controls were analyzed using Fisher's exact test. Results The genotype and allele frequencies of the C1019T polymorphism significantly differ between cases and controls. The frequencies of C/C genotype ( P = 0.009) and ‘ C ’ allele ( P = 0.002) of the C1019T polymorphism showed a significant prevalence in cases compared to controls. Conclusion Our findings suggest that the Cx37 C1019T variation may contribute to the risk of PCOS in the South Indian women.

Development of non-invasive steroid hormone assays using fecal samples is crucial for detection of pregnancy and monitoring of fertility status in big cats and thus facilitates conservation and management of wild animals. Due to changes in metabolism and excretory pattern, animals excrete different steroid metabolites in feces and urine. The present study is an attempt to develop a common enzyme immunoassay for 5α-pregnan-3α-ol-20-one one of the predominant progestogen metabolites in the feces samples of big cats. The developed ELISA showed a high sensitivity and low cross reactivity to other hormones compared to commercially available RIA kits based on progesterone antibody. It could be used in a wide range of animals for monitoring fertility status and pregnancy detection by measuring fecal steroid metabolites.

Transrectal ultrasonography was used to monitor the ovarian follicular dynamics in the Indian blackbuck antelope ( Antilope cervicapra ). The blackbuck has a short oestrous cycle (mean length 15.3 ± 0.8 days) and exhibits two follicular waves, with a mean time of 14.3 ± 0.8 and 8.9 ± 0.6 days for the first and second waves, respectively. The mean diameter of the preovulatory follicles was 0.54 ± 0.04 cm, and only a single ovulatory follicle resulted from the second follicular wave in all the hinds. Follicular activity and preovulatory follicles appeared alternatively on the ovaries in successive cycles. The plasma estradiol-17β (E 2 ) concentrations also exhibited a wave-like pattern, varying between 1.3 ± 0.01 pg ml −1 and 5.3 ± 1.9 pg ml −1 . A positive relationship ( r = 0.85) was recorded between the dominant follicle diameter and the plasma E 2 concentrations during the oestrous cycle. Furthermore, this baseline ultrasound data, in particular the diameter of the preovulatory follicle, enabled the team to carry out fixed-time AI, leading to live offspring in this native threatened species. This study, for the first time, documents the use of transrectal ultrasonography to monitor ovarian follicular dynamics in the Indian blackbuck antelope. It was also successful in the application and the development of a protocol for fixed-time AI in the Indian blackbuck antelope.

The aim of the study was to evaluate meiotic maturation, and expression of genes coding for oocyte secreted factors (GDF9, BMP15, TGFBR1, and BPR2) and apoptosis (BCL2, BAX and P53) after vitrification of immature goat cumulus oocyte complexes (COCs) and in vitro maturation. COCs were vitrified in a solution containing ethylene glycol, dimethyl sulfoxide and sucrose using either a conventional straw (CS), open pulled straw (OPS), cryoloop (CL), hemistraw (HS) or cryotop (CT). Freshly collected COCs (Control), COCs exposed to vitrification and dilution solutions without cryopreservation (EC) and vitrified-warmed COCs were matured in vitro for 27h. The viability of vitrified-warmed COCs 2 h post warming and in vitro maturation was similar for CL, HS and CT. The proportion of oocytes that extruded a 1st polar body and reached TI/MII was significantly higher with CT and HS followed by CL, OPS and CS. Gene expression of GDF9, BMP15, BMPR2, BAX and P53 were comparable to control levels for OPS, CL, HS and CT. The gene expression pattern in CS vitrified COCs was by contrast changed in that GDF9, BMP15, TGFBR1 and BAX were up regulated and BMPR2, BCL2 and P53 down regulated. In conclusion immature goat COCs vitrified using CT and HS showed that viability, maturation rates and expression of genes coding for oocyte secreted factors and apoptosis were similar to non-vitrified controls.

Hepatozoon species are parasites that infect a wide variety of domestic and wild animals. The objective of this study was to perform the molecular detection and characterization of Hepatozoon spp. in Asiatic lion, Indian tiger, Indian leopard, Indian wild dog, Indian domestic dog and cat based on partial 18S rRNA gene sequences from Hepatozoon spp. in the naturally infected animals. Hepatozoon spp. could be detected in blood samples of 5 out of 9 Asiatic lions, 2 out of 5 Indian tigers, 2 out of 4 Indian leopards and 2 out of 2 Indian wild dogs and, 2 out of 4 domestic cats and 2 out of 3 domestic dog samples by PCR. Sequencing of PCR amplicon and BLAST analysis of partial 18S rRNA gene sequences indicated that the Hepatozoon spp. in Asiatic lion, Bengal tiger, Indian leopard and domestic cat was Hepatozoon felis (98-99% similarity) and in the Indian wild and domestic dog the phylogenetic neighbour was Hepatozoon canis (97-100% similarity). Presence of H. felis and H. canis in both domestic and wild animals suggested that they are not host specific and the same parasite causes infection in domestic and wild felids and canids in India and from different parts of the world. To our knowledge, this is the first report on detection and molecular characterization of H. felis infection in Asiatic lions, Indian tigers, Indian leopards and H. canis in Indian wild dog. Hepatozoon spp. may be a potential pathogen and an opportunistic parasite in immuno-compromised animals and could thus represent a threat to endangered Indian wild felids and canids.

Hepatozoon species are parasites that infect a wide variety of domestic and wild animals. The objective of the study was to detect the occurrence of Hepatozoon ursi in Indian sloth bears and to characterize the parasite based on phylogenetic analysis of the partial 18S rRNA gene sequence. Hepatozoon infection could be detected in 38 (70%) out of fifty-four blood samples of Indian sloth bears (captive and wild), suggestive of high prevalence of Hepatozoon infection in Indian sloth bears. Sequencing of partial 18S rRNA gene of the positive samples and BLAST analysis indicated that the nearest phylogenetic neighbour was H. ursi with which they exhibited 99-100% similarity. Additionally, Hepatozoon sp. isolated from wild sloth bears of India were identical to those in captive sloth bears and phylogenetically related to H. ursi reported from Japanese black bears from Japan. To our knowledge, this is the first report on the molecular characterization of H. ursi infection in Indian sloth bears.

Cell-free culture supernatants of five psychrophilic bacteria Pseudomonas antarctica, Pseudomonas proteolytica, Pseudomonas meridiana, Arthrobacter kerguelensis and Arthrobacter gangotriensis and two mesophilic bacteria Bacillus indicus and Bacillus cecembensis have been used to synthesize silver nanoparticles (AgNPs). The AgNPs were characterized using UV–Visible spectroscopy, transmission electron microscopy and atomic force microscopy. The sizes of the AgNPs ranged from 6 to 13 nm and were stable for 8 months in the dark. The synthesis and stability of AgNPs appears to depend on the temperature, pH or the species of bacteria (P. antarctica or A. kerguelensis) from which the supernatant is used. It was observed that the A. kerguelensis supernatant could not produce AgNPs at the temperature where the P. antarctica could synthesize AgNPs. Thus the study provides evidence that the factors in the cell-free culture supernatants that facilitate synthesis of AgNPs vary from bacterial species to species. The AgNPs were bacteriocidal. This report presents for the first time data on the generation of AgNPs using culture supernatants of psychrophilic bacteria and also demonstrates that culture supernatants of species of Arthrobacter, a genus which has never been investigated earlier, can also synthesize AgNPs.

Bacterial diversity of the subsurface (18–22 cm), middle (60–64 cm) and bottom (100–104 cm) of a 136-cm-long sediment core sampled from a freshwater lake in Antarctica was determined by the culturable approach, T-RFLP and 16S rRNA gene clone libraries. Using the culturable approach, 41 strains were isolated and, based on phylogenetic analysis, they could be categorized into 14 groups. Representatives of the 14 groups varied in their growth temperature range (4–30 °C), in their tolerance to NaCl (0–2 M NaCl) and in the growth pH range (5–11). Eleven of fourteen representative strains exhibited either amylase, lipase, protease and (or) urease activities at 4 °C. Bacterial diversity at the phyla level using T-RFLP and 16S rRNA clone libraries was similar and clones were affiliated with Proteobacteria, Bacteroidetes, Actinobacteria and Firmicutes . TRFs affiliated with Spirochaetes were detected only by the T-RFLP approach and clones affiliated with Caldiserica only in the clone libraries. Stratification of bacteria along the depth of the sediment was observed both with the T-RFLP and the 16S rRNA gene clone library methods, and results indicated that stratification was dependent on the nature of the organism, aerobic or anaerobic. For instance, aerobic Janthinobacterium and Polaromonas were confined to the surface of the sediment, whereas anaerobic Caldisericum was present only in the bottom portion of the core. It may be concluded that the bacterial diversity of an Antarctic lake sediment core sample varies throughout the length of the core depending on the oxic–anoxic conditions of the sediment. Furthermore, these psychrophilic bacteria, due to their ability to produce extracellular cold active enzymes, might play a key role in the transformation of complex organic compounds.

A cold-active heat-labile t-RNA modification GTPase (TrmE) from psychrophilic bacterium Pseudomonas syringae (Lz4W) has been purified and characterized. The purified TrmE is a 53 kDa protein, has GTPase activity and hydrolyses only the oxy and deoxy forms of GTP but not the other nucleotide triphosphates. The enzyme exhibits optimal activity at 12-18 degrees C and retains 65% of its optimal activity at 4 degrees C, indicating that it is a cold-active enzyme. The enzyme is also heat-labile and loses 60% of its activity at 30 degrees C. This is the first report on the purification and characterization of a TrmE from a psychrophilic bacterium.

Spermatozoa deposited in the female reproductive tract need to undergo a multifaceted maturation process prior to fertilization termed "capacitation". This process is regulated by several proteins which are compartmentalized in discrete domains of the spermatozoon including the head, the mid-piece and the principal piece. Over the last decade many proteins involved in capacitation have been identified, such as proteins involved in the organization of the tail, proteins involved in signal transduction, chaperones, ion-channel proteins and mitochondria-associated proteins. This review focuses on the identity and function of mitochondrial proteins which undergo capacitation-dependent tyrosine phosphorylation in spermatozoa.

Culturable bacterial diversity of Midtre Lo v ´ enbreen glacier, an Arctic glacier, was studied using 12 sediment samples collected from different points, along a transect, from the snout of Midtre Lo v ´ enbreen glacier up to the convergence point of the melt water stream with the sea. Bacterial abundance appeared to be closer to the convergence point of the glacial melt water stream with the sea than at the snout of the glacier. A total of 117 bacterial strains were isolated from the sediment samples. Based on 16S rRNA gene sequence analyses, the isolates ( n = 117) could be categorised in to 32 groups, with each group representing a different taxa belonging to 4 phyla ( Actinobacteria , Bacilli , Flavobacteria and Proteobacteria ). Representatives of the 32 groups varied in their growth temperature range (4–37 °C), in their tolerance to NaCl (0.1–1 M NaCl) and in the growth pH range (2–13). Only 14 of 32 representative strains exhibited amylase, lipase and (or) protease activity and only one isolate (AsdM4-6) showed all three enzyme activities at 5 and 20 °C respectively. More than half of the isolates were pigmented. Fatty acid profile studies indicated that short-chain fatty acids, unsaturated fatty acids, branched fatty acids, cyclic and cis fatty acids are predominant in the psychrophilic bacteria .

Objective To determine the effectiveness of yohimbine as an antagonist of ketamine-xylazine anaesthesia in captive Asiatic lions ( Panthera leo persica ), tigers ( Panthera tigris ) and leopards ( Panthera pardus ). Study design Prospective clinical trial. Animals Fifty-two healthy adult lions, 55 adult leopards and 16 adult male tigers. Methods Captive wild felids in Indian zoos were anaesthetized with a combination of ketamine (2.2-2.6 mg kg −1 ) and xylazine (1.1-1.3 mg kg −1 ) using a dart propelled from a blowpipe. Time to onset of anaesthesia, lateral recumbency and induction time were measured, and physiological variables (respiration, heart rate and rectal temperature) were recorded once after the onset of complete anaesthesia. Anaesthesia was antagonized at various time periods with an intravenous administration of either 0.1 or 0.15 mg kg −1 yohimbine. Onset of arousal and time to complete anaesthetic recovery were recorded. Results A total of 123 immobilizations were conducted between 2000 and 2005. Anaesthetic induction was achieved in 15-25 minutes in all animals. Incidents of sudden recovery or life-threatening effects associated with immobilizations were not observed. Yohimbine effectively antagonized anaesthesia in all animals within 10 minutes without any excitatory behaviour compared to control animals. No adverse reactions/side effects to yohimbine were recorded except that a few leopards exhibited seizure-like signs for a short period immediately after yohimbine administration. The duration of anaesthesia had no significant effect on the recovery time in any of the animals. Conclusion and clinical relevance Yohimbine antagonized the xylazine portion of ketamine-xylazine anaesthesia and thereby hastened recovery from anaesthesia in Asiatic lions, tigers and leopards.

Objective: To investigate whether the eNOS gene influences the risk of developing endometriosis in south Indian women. Study design: The single nucleotide polymorphism, Glu298Asp, in exon7 of the eNOS gene was tested for association in a case-control study of 232 affected women and 210 women with no evidence of disease. All the women were infertile, ascertained from the same infertility clinic. The genotype frequencies of the polymorphism were compared, using polymerase chain reaction and sequencing analysis. The localization and expression of eNOS in the eutopic endometrium of five cases and four controls was also analyzed using immunohistochemistry and western blotting. Results: No statistically significant differences were observed in the genotype distributions and allele frequencies (p = 0.3) between the cases and controls according to codominant, dominant and recessive genotype models. The localization and expression of this protein were similar in the endometrium of cases and controls. Conclusion: In the present study we could neither observe a difference in the eNOS expression nor establish an association between the eNOS Glu298Asp exon 7 polymorphism in south Indian women with endometriosis.

In this paper the behavior of the Asiatic lion was studied during natural and gonadotropin-induced estrus in relation to fecal estradiol and progesterone concentration. The average length of estrus was 5.4 days and no significant difference was observed between natural and gonadotropin-induced estrus. Vocalization and rolling were the major estrus behavioral activities of Asiatic lions and the frequency of these activities were similar in both natural and gonadotropin-induced estrus and treatment with exogenous gonadotropin did not alter estrus behavioral activities. A significant positive correlation was observed between fecal estradiol and frequency of estrus behavior during natural and gonadotropin-induced estrus. Following gonadotropin treatment estrus could be induced in 69% of animals and these induced animals ovulated following hCG treatment. This study reports for the first time the successful use of the non-invasive fecal steroid assay for monitoring the induction of estrus and ovulation in the Asiatic lion.

Ten adult male spotted deer were monitored over a 2-year interval to determine seasonal variations in testicular size, semen characteristics and serum testosterone concentrations, and to determine if there was an association between season and type of antler. Mean (+/-S.E.M.) testicular volume (118.8+/-4.6 cm(3)), serum testosterone concentration (1.2+/-0.1 ng/mL), semen volume (4.1+/-0.6 mL), sperm concentration (338.3+/-24.9 x 10(6) mL(-1)), percentage of morphologically normal sperm (79.1+/-2.8%), and percentage of motile sperm (66.5+/-1.5%) were higher (P

To be fertilization competent, spermatozoa undergo a series of changes in the female reproductive tract collectively referred to as capacitation. In an attempt to understand, if ornidazole, a known anti-fertility drug, adversely affects sperm functions by targeting capacitation, we designed experiments to study the influence of this drug on hyperactivation (HA), capacitation-associated protein tyrosine phosphorylation (pY) and the acrosome reaction (AR). Addition of ornidazole at 0 h, inhibited the onset of HA and total pY in a dose dependent manner. However, when ornidazole was added at 3.5h, severe effects were still seen on HA and pY of high molecular weight proteins but, pY of lower M(r) proteins (50-56 kDa) was affected only marginally. Further, lower doses of ornidazole (5 and 10 mM) had greater inhibitory effect when added at 0 h, while addition of ornidazole at 3.5 h required higher doses of ornidazole (25 mM) to cause significant inhibition of acrosome reaction. Collectively, through in vitro studies, we demonstrate that ornidazole affects the onset and progression of hamster sperm hyperactivation, capacitation associated protein tyrosine phosphorylation and acrosome reaction, and the severity depends on the dose (5, 10 or 25 mM) and the time of addition (0 or 3.5 h) of the drug to the spermatozoa.

Spermatozoa are cells distinctly different from other somatic cells of the body, capacitation being one of the unique phenomena manifested by this gamete. We have shown earlier that dihydrolipoamide dehydrogenase, a post-pyruvate metabolic enzyme, undergoes capacitation-dependent tyrosine phosphorylation, and the functioning of the enzyme is required for hyperactivation (enhanced motility) and acrosome reaction of hamster spermatozoa (Mitra, K., and Shivaji, S. (2004) Biol. Reprod. 70, 887-899). In this report we have investigated the localization of this mitochondrial enzyme in spermatozoa revealing non-canonical extra-mitochondrial localization of the enzyme in mammalian spermatozoa. In hamster spermatozoa, dihydrolipoamide dehydrogenase along with its host complex, the pyruvate dehydrogenase complex, are localized in the acrosome and in the principal piece of the sperm flagella. The localization of dihydrolipoamide dehydrogenase, however, appears to be in the mitochondria in the spermatocytes, but in spermatids it appears to show a juxtanuclear localization (like Golgi). The capacitation-dependent time course of tyrosine phosphorylation of dihydrolipoamide dehydrogenase appears to be different in the principal piece of the flagella and the acrosome in hamster spermatozoa. Activity assays of this bi-directional enzyme suggest a strong correlation between the tyrosine phosphorylation and the bi-directional enzyme activity. This is the first report of a direct correlation of the localization, tyrosine phosphorylation, and activity of the important metabolic enzyme, dihydrolipoamide dehydrogenase, implicating dual involvement and regulation of the enzyme during sperm capacitation.

One anaerobic, proteolytic bacterium isolated from Schirmacher oasis, Antarctica was characterised taxonomically. Based on morphology, biochemical characteristics and 16S rRNA sequence the isolate was identified as Clostridium species with closest similarity with Clostridium subterminale. Isolate was psychrotrophic forming maximum cell mass between 5 and 10 °C and produced extracellular protease. Growth was observed in the pH range of 6.5-8.5 with optimum at pH 8. Protease was purified 12.7-fold with a total yield of 26.2%. Effect of temperature, pH and salt concentration on enzyme activity were studied. Protease was found to be a serine-type metaloenzyme, which is active in a broad range of pH. It was moderately thermolabile and resistant to SDS. Enzyme kinetics revealed a tendency to decrease K m with increase in temperature for casein substrate.

Nine psychrotolerant bacteria were isolated from fast ice in the middle of Geologie Archipelago, Adelie Land, Antarctica and were categorized into two groups, based on their SDS-PAGE profiles. Representatives from each of the two groups, namely strains DD 48 T and SJ 14 T exhibited phenotypic and chemotaxonomic characteristics confirming to the genus Psychrobacter. The 16S rRNA gene sequence indicated that the two isolates are closely related to each other and to the already reported fifteen species of Psychrobacter. Detailed studies on the phenotypic characteristics, chemotaxonomic properties and phylogenetic analysis of strains DD 48 T and SJ 14 T indicated that they are distinctly different from each other and the reported species of Psychrobacter. At the DNA-DNA hybridisation level, the two species exhibit less than 70% similarity. Thus, strains DD 48 T and SJ 14 T are identified as new species of the genus Psychrobacter for which the names Psychrobacter salsus sp. nov. and Psychrobacter adeliensis sp. nov. respectively are proposed.

We report the 4.86-Mb draft genome sequence of Bacillus okhensis strain Kh10-101T, a halo-alkali tolerant rod shaped bacterium isolated from a salt pan near port of Okha, India. This bacterium is a potential model to study the molecular response of bacteria to salt as well as alkaline stress, as it thrives under both high salt and high pH conditions. The draft genome consist of 4,865,284bp with 38.2% G+C, 4952 predicted CDS, 157 tRNAs and 8 rRNAs. Sequence was deposited at DDBJ/EMBL/GenBank under the project accession JRJU00000000.

The carotenoid composition of a psychrotrophic bacterium Micrococcus roseus strain MTCC 678 has been re-examined qualitatively and quantitatively by chromatographic (TLC, HPLC) and spectroscopic (VIS, MS, 1H NMR and CD) methods, partly in direct comparison with authentic carotenoids. The carotenoid content of 0.13% of the dry weight consisted of (2S,2′S)-bactedorubedn (68% of total), bacterioruberin monoglycoside (21%), monoanhydrobacterioruberin (6%), an unidentified polar tridecasne carotenoid (3%), haloxanthin (2%) and β,β-carotene (0.4%): a carotenoid complement associated with that of halophilic bacteria. Rapid analysis demonstrated that bacterioruberin occurred as the all-E isomer (75% of total), together with the 5Z-(5%), 9Z-(5%) and 13Z-(15%) geometrical isomers. The established interaction of the major carotenoid with membrane vesicles, compatible with cold adaptation, is rationalized.

Photoacoustic spectra of cells of M. roseus were recorded to study the in vivo characteristics and localisation of the carotenoid pigments in these cells. The PA spectra indicated that both the psychrotrophic and mesophilic strains had similar chromophores. The cis carotenoids were prominent in the psychrotrophic M.roseus whereas shorter polyenes were more prominent in mesophilic M.roseus. Further, depth profiling photoacoustic studies revealed that in both the strains of M.roseus the bulk of the chromophore was associated with the cell membrane.

Summary Two acidophilic, aerobic, heterotrophic, gram-negative, nonsporulating bacterial strains which were isolated from an acidic copper mine, possessed the unusual capacity to grow at a pH above 6.0 in a mineral salt-glucose-yeast extract medium. Results of 16S rDNA analysis revealed that strain GS 18 h shares high sequence similarity with Acidiphilium cryptum and “ Acidiphilium symbioticum ” (both 99.9% homology) as well as with Acidiphilium organovorum (99.5% homology). Similarity values obtained from DNA-DNA hybridization studies between strain GS 18 h and A. cryptum and “ A. symbioticum ” indicated the lack of relatedness at the strain level (40–51% similarity). The sequence of the 16S rDNA of strain GS 19 h was found to be related to that of Acidiphilium aminolyticum (99.1% similarity), a species that has recently been transferred to Acidocella gen. nov. as Acidocella aminolytica .

The fluorescence excitation and emission spectra are reported for P-3 (bis-dehydro-B-carotene-2-carboxylic acid), the major carotenoid pigment of psychrotrophic M. roseus. The excitation spectrum and the absorption spectrum showed good agreement with respect to the position of their peak maxima. The study also demonstrates that P-3 binds to liposomes prepared from synthetic lipids (PC, DOPG, or CL) or the total lipids of a mutant colourless M. roseus. Binding of P-3 to the membranes was accompanied by a decrease in the fluorescence emission intensity and a blue shift in lambda(em) maximum by 15 to 20 nm. The quantum yield of P-3 was observed to be low (1.7 x 10(-5)).

The interaction of progesterone, 17-α-hydroxyprogesterone, testosterone and estradiol with membrane vesicles prepared from phosphatidylserine (PS), from the total lipids of human and hamster spermatozoa, from the lipids of hamster spermatozoal plasma and acrosomal membrane and with the native membranes of hamster spermatozoa have been investigated by 90° light scattering and fluorescence spectroscopy. The results indicate that progesterone decreases the fluidity of membranes, aggregates membrane vesicles, induces fusiionn of membrane vesicles and also renders them permeable to hydrophilic molecules like car☐yfluorescein. But, testosterone and estradiol at the same concentration had very little effect on membrane fluidity, membrane aggregation, fusion and leakage. The above membrane perturbing activities of the steroids is discussed in light of the recent findings that progesterone induces acrosome reaction in human and hamster spermatozoa [11, 18].

Summary An age dependent study of RNA and protein syntheses was carried out in S. notophora using radioactive precursors. RNA synthesis in the ciliate steadily increased from G 1 phase and showed a peak at the end of S phase (8 h) while protein synthesis showed a peak at 6 h. 3 H-lysine incorporation was found to be more than leucine. The effect of metepa on the synthesis of RNA and proteins was also studied by treating ciliates in different phases of the cell cycle with different concentrations of metepa. Treatment reduced the synthesis of RNA and proteins in the G 1 , S and G 2 phases as compared to controls. But in general the S phase ciliates were the most susceptible and the G 2 phase were the least affected. Higher concentrations of metepa had an increased inhibitory affect on RNA and protein synthesis. However, the treated ciliates showed increase in synthesis of RNA and proteins when allowed to recover.

Semen, like blood, has two types of components — the cellular spermatozoa and the non-cellular seminal plasma. Seminal plasma contains the combined secretion of all the male accessory sex glands and is of great physiological significance since it serves as the carrier of sperm to the female reproductive tract; it also influences sperm motility and the acrosome reaction. These physiological attributes of seminal plasma have been widely discussed but the antimicrobial activity of seminal plasma remains relatively unknown.

Changes in the cell surface of vaginal epithelial cells were studied by scanning microscopy and fluorescence spectroscopy. Microvilli which are prominent features of the vaginal epithelial cells in proestrus and diestrus are replaced by sheet-like structures in the estrus phase. Surface morphology of vaginal epithelial cells of estradiol primed rat resembles the vaginal cells from estrus phase rats whereas vaginal cells from control rats resembles the diestrus phase. Measurement of the fluidity of the membranes indicated that the vaginal epithelial cell membrane of estrus rats is more fluid compared to proestrus and diestrus. Similarly, estradiol primed immature rat vaginal epithelial cell membrane was observed to be more fluid than the corresponding control.