Heat shock protein 70 expression in different tissues of Cirrhinus mrigala (Ham.) following heat stress

Members of heat shock protein 70 (HSP70) are highly conserved proteins of about 70 kDa and play important roles in protein folding. Levels of these proteins increase when cells are under stress. Environmental temperature influences both the basal and induced levels of HSPs. However, studies on HSPs in fishes from a tropical country such as India are lacking. In the present study, Indian major carp (IMC) Cirrhinus mrigala (Ham.) acclimatized at 25±2°C had high levels of HSP70, viz., 1.2–1.3 ng μg^?1 total protein in kidney and gill and 4.2–5.3 ng μg^?1 total protein in liver and brain tissues, indicating the presence of biochemically significant levels of stress. However, maintenance of acclimatized fish at 17°C for up to 48 h did not lead to a significant decrease in stress protein levels. A heat shock at 37°C for up to 48 h resulted in only two to threefold increase in HSP70 levels in these organs. Although the increase in HSP70 levels was apparent from the first hour of heat stress in all these tissues, the increase was significant from the second hour in the brain, the sixth hour in liver and kidney and the 20th hour in the gills.     

Nitrous oxide emission measurement with acetylene inhibition method in paddy fields under flood conditions

Nitrous oxide (N₂O) emission from flooded rice paddy fields was continuously measured by the closed chamber method at an experimental plot in Thailand for a whole cultivation period. To characterize the N₂O emission with regard to the denitrification N loss, the C₂H₂ inhibition method was applied. Flood water on the soil greatly suppressed the N₂O emission. The N₂O emission was mitigated considerably by even a thin film of the flood water. The overall average N₂O emissions under flood conditions for one crop season (83 days) at the control site and the C₂H₂ treated site were 10.3 and 11.8 μg N m⁻² h⁻¹, respectively. The N₂O emission from the C₂H₂ treated site was consistently higher than that from the control site and the N₂O emission from both sites followed the same diurnal and seasonal variation pattern, indicating the effect of denitrification inhibition by the supplied C₂H₂. The N₂O emission enhanced along with temperature increase when NO₃–N concentration in the soil water was above 0.4 mg N l⁻¹ and soil temperature was above 24°C, suggesting specific temperature influence over the emission. The increase in NO₃–N concentration and temperature in the soil affected only the N₂O emission while the difference in the emission at the C₂H₂ treated site and the control site was not so much affected. It was suggested that most of the actively produced N₂O under higher NO₃–N concentration and temperature would likely to quickly emit to the atmosphere rather than to undergo further reduction to N₂.     

Priority habitats for the conservation of large river fish in the Ganges river basin

• 1. Three classes of habitat used by groups of fish species classified as conservation and management priorities were developed for the Gerua River (also known as the Girwa River, Karnali River) in the Ganges river basin. This river is large (mean annual discharge ca 1500 m³ s^?1, up to 900 m wide), surrounded by protected lands of India and Nepal, and upstream of major diversions and river alterations.
• 2. Fish and habitat sampling was conducted at 45 sites from 2000 to 2003. Data were analysed for 2172 fish of 14 species. Species and life stages found occupying a statistically distinct subset of the river habitats were grouped to identify classes of river habitat for conservation.
• 3. Most species and life‐stage groups specialized on specific habitat conditions revealed by multivariate analyses of variance and a principal component analysis. The most numerous and diverse group (six species, 15 life stages) was associated with deep depositional habitats with sandy substrate. Two species covering three life stages were primarily oriented to erosional habitat marked by fast current velocity with pebble and cobble substrate. A third group of three species of adults and juveniles were intermediate in habitat use.
• 4. River conservation for fish faunas should maintain both erosional and depositional channel habitats with depths, substrates, and current velocity inclusive of the ranges reported. The erosional and depositional nature of the key habitats requires that rivers be maintained with flows capable of channel‐forming functions.

Copyright © 2006 John Wiley & Sons, Ltd.     

Effect of supplementing vitamin E analogues on post-thaw semen parameters and fertility in chicken

1. The effect of supplementing water-soluble vitamin E analogues 6-hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid (trolox) and butylated hydroxy toluene (BHT) was studied in two separate experiments.

2. In the first experiment, trolox was supplemented at 0.2 mM, 0.4 mM and 0.8 mM concentrations along with N-methylacetamide (MA; 12% final concentration) and semen was cryopreserved in 0.5 ml French straws. Different semen parameters and fertility were assessed from post-thaw samples.

3. Sperm motility, live sperm, and mitochondrial activity were significantly lower (P < 0.05) in cryopreserved semen. Lipid peroxidation (LPO) was significantly higher (P < 0.05) in cryopreserved semen that was reduced by trolox supplementation. The treatment containing trolox at 0.2 mM concentration produced significantly higher (P < 0.05) fertility compared to unsupplemented cryopreservation treatment.

4. In the second experiment, BHT was supplemented at 0.25 mM, 0.5 mM, and 1 mM concentrations along with MA during semen cryopreservation.

5. Sperm motility, live sperm and MTT dye reduction test were significantly lower (P < 0.05) in cryopreserved semen. These parameters declined with increasing BHT concentration. Abnormal sperm was significantly higher (P < 0.05) in the BHT supplemented treatments. The sperm chromatin dispersion (SCD) test was significantly higher (P < 0.05) in cryopreserved samples and was highest in samples supplemented with 0.5 mM and 1 mM BHT. The percentage fertility was significantly lower (P < 0.05) in cryopreserved semen and BHT supplementation did not improve fertility.

6. In conclusion, trolox supplementation at 0.2 mM concentration during semen cryopreservation improved fertility, whereas BHT supplementation resulted in a decline in post-thaw semen parameters.     

Association of bovine KISS1 single nucleotide polymorphisms with reproductive traits in Indian Cattle

Kisspeptins, a family of neuropeptide encoded by the Kiss1 gene, have emerged as crucial regulator of fertility and reproduction by regulating the hypothalamic–pituitary–gonadal axis. The present study was aimed to identify and associate SNPs in the KISS1 gene with reproductive traits in cattle of Indian origin. DNA samples collected from 300 individual cows of three Indian dairy breeds (Gir, Kankrej and Frieswal) of cattle were used in the study. The SNPs of KISS1 gene were identified with PCR-RFLP and sequence analysis using two sets of primer pairs. A total of 5 SNPs were identified in the targeted region of which, two were selected for screening the population and association studies. The analysis revealed that genotypes of rs442633552G>A and rs42022871C>T had a significant association with dry period. The SNP rs42022871C>T also established significant role in milk production traits, and selection of TT-genotyped animals will improve the reproduction and production potential of the animals.     

Temporal changes in endogenous estrogens and expression of behaviors associated with estrus during the periovulaory period in Murrah buffaloes (<Emphasis Type="Italic">Bubalus bubalis</Emphasis>)

The objective of this study were (1) to establish the duration of behavioral estrus signs and timing of ovulation in Murrah buffaloes (n = 10) and (2) to determine relationship between behavioral estrus signs with change in plasma estrogen concentrations. Estrus and its behavioral signs were detected at hourly intervals by visual observations, per recta examination of genitalia and bull parading four times in a day for 30 minutes each. Among the behavioral signs of estrus, swollen vulva (80%) was the best indicator of estrus followed by excitement (70%). Among the duration of behavioral estrus signs the first and longest duration of estrus signs was swollen vulva which was seen upto 19.8 ± 0.8 h after onset of estrus. The mean total duration of estrus symptoms from appearance to disappearance of all the behavioral estrus symptoms was 23.5 ± 1.7 h. All the behavioral estrus symptoms were observed during the period of estrogen surge. Endocrine profile during the periestrus period showed that the mean peak concentrations of total estrogen 23.9 ± 3.9 pg/ml occurred at 8.8 ± 1.7 h after onset of estrus. The average number of estrus symptoms observed per animal during onset of spontaneous estrus was 5.7. Ovulation occurred after 37.4 ± 1.7 h after onset of estrus and 13.4 ± 1.0 h after end of total estrogen surge respectively. In conclusion, our results suggest that all signs of behavioral estrus occurred during the preovulatory rise in estrogens. The first sign of estrus to be observed was a swollen vulva and this symptom persisted the longest.     

Experimental avian paramyxovirus serotype-3 infection in chickens and turkeys

Avian paramyxoviruses (APMV) are divided into nine serotypes. Newcastle disease virus (APMV-1) is the most extensively characterized, while relatively little information is available for the other APMV serotypes. In the present study, we examined the pathogenicity of two divergent strains of APMV-3, Netherlands and Wisconsin, in (i) 9-day-old embryonated chicken eggs, (ii) 1-day-old specific pathogen free (SPF) chicks and turkeys, and (iii) 2-week-old SPF chickens and turkeys. The mean death time in 9-day-old embryonated chicken eggs was 112 h for APMV-3 strain Netherlands and > 168 h for strain Wisconsin. The intracerebral pathogenicity index in 1-day-old chicks for strain Netherlands was 0.39 and for strain Wisconsin was zero. Thus, both strains are lentogenic. Both the strains replicated well in brain tissue when inoculated intracerebrally in 1-day-old SPF chicks, but without causing death. Mild respiratory disease signs were observed in 1-day-old chickens and turkeys when inoculated through oculonasal route with either strain. There were no overt signs of illness in 2-weeks-old chickens and turkeys by either strain, although all the birds seroconverted after infection. The viruses were isolated predominantly from brain, lungs, spleens, trachea, pancreas and kidney. Immunohistochemistry studies also showed the presence of large amount of viral antigens in both epithelial and sub-epithelial lining of respiratory and alimentary tracts. Our result suggests systemic spread of APMV-3 even though the viral fusion glycoprotein does not contain the canonical furin proteases cleavage site. Furthermore, there was little or no disease despite systemic viral spread and abundant viral replication in all the tissues tested.     

Identification of unique DNA sequences present in highly virulent 2009 Alabama isolates of Aeromonas hydrophila

In 2009, a disease outbreak caused by Aeromonas hydrophila occurred in 48 catfish farms in West Alabama, causing an estimated loss of more than 3 million pounds of food size channel catfish. Virulence studies have revealed that the 2009 isolates of A. hydrophila are at least 200-fold more virulent than a 1998 Alabama isolate AL98-C1B. However, up to now, no molecular markers have been identified to differentiate the highly virulent 2009 isolates from other isolates of A. hydrophila. To understand the genetic differences between the highly virulent 2009 isolates and the less virulent AL98-C1B at molecular level, PCR-select bacterial genome subtractive hybridization was used in this study. A total of 96 clones were selected from the subtractive genomic DNA library. Sequencing results revealed that the 96 clones represented 64 unique A. hydrophila sequences. Of the 64 sequences, three (hypothetical protein XAUC_13870, structural toxin protein RtxA, and putative methyltransferase) were confirmed to be present in the three virulent 2009 Alabama isolates but absent in the less virulent AL98-C1B. Using genomic DNAs from nine field isolates of A. hydrophila with different virulence as templates, two sequences (hypothetical protein XAUC_13870 and putative methyltransferase) were found to be only present in highly virulent A. hydrophila isolates, but absent in avirulent isolates.     

Prevalence and duration of insensibility following electrical stunning in calves

Sixty-one calves were electrically stunned with 100-250 V currents (103-1806 mA) and the prevalence and duration of insensibility was assessed from their physical behaviour, the presence of epileptiform activity in their electrocorticograms, and the absence of visual evoked responses in their electrocorticograms. A current applied at 100 V across the head for 3 s failed to induce insensibility in all cases. Currents at 150-250 V induced insensibility in all calves and the shortest duration of insensibility was 44 s. It is recommended that a 150-200 V current would be appropriate for commercial use.