A multiplex PCR assay to detect mislabelling in fish products.

Fish substitution in fish products is an important issue in fish markets, as it is a widespread practice. An authentication protocol for Rohu, Thaila and Tilapia was developed by multiplex PCR. Three species-specific and one degenerate common forward primer were designed using the Cytb gene of the mitochondrial genome. These primers for Labeo rohita, Labeo catla and Oreochromis niloticus showed the fragment size of 235 bp, 186 bp and 506 bp on the agarose gel, respectively. The primers for L. rohita and L. catla were sensitive to 0.1 ng of DNA template, while for O. niloticus this value was 1 ng of DNA template. A total of 230 commercial samples (160 fried and 70 processed fish products) were screened, where 60% mislabeling in fried and 30% mislabeling in processed fish were found. This multiplex PCR protocol could give useful insights for food inspection and enforcement of regulatory food control.

Diversity and genetic structure of freshwater shark Wallago attu: an emerging species of commercial interest.

Pakistan has natural freshwater resources acting as a hotspot for diverse fish fauna. However, this aquatic fauna is declining at an alarming rate due to over-exploitation, habitat degradation, water pollution, climate change, and certain anthropogenic activities. The freshwater shark, Wallago attu, is a popular edible catfish inhabiting these freshwater ecosystems. Habitat degradation, overfishing, and human activities are heavily impacting the natural population of this species. So, sound knowledge about its population structure is necessary for its proper management in natural waters. The current study involves utilizing two mtDNA markers (COI, Cytb) to assess the genetic structure and differentiation among W. attu populations of Pakistani Rivers. Genetic variability analysis indicated a high haplotype (0.343 ± 0.046-0.870 ± 0.023) and low nucleotide diversity (0.0024 ± 0.012-0.0038 ± 0.018) among single and combined gene sequences, respectively. Overall, River Indus was populated with more diverse fauna of Wallago attu as compared to River Chenab and River Ravi. Population pairwise, Fst values (0.40-0.61) were found to be significantly different (p < 0.01) among three Riverine populations based upon combined gene sequences. The gene flow for the combined gene (COI + Cytb) dataset among three populations was less than 1.0. The transition/transversion bias value R (0.58) was calculated for testing of neutral evolution, and it declared low genetic polymorphism among natural riverine populations of Wallago attu. The current study's findings would be meaningful in planning the management and conservation of this economically important catfish in future.

Geographic distribution and genetic diversity of Newcastle disease virus in pigeons from Pakistan.

RESEARCH HIGHLIGHTS: Virulent NDV genotypes were repeatedly isolated from pigeons.Evidence of epidemiological links among viruses isolated from various locations.Distinct phylogenetic branches suggest separate, simultaneous evolution of NDVs.Study information could be helpful in the development of an effective vaccine.

Relative expression levels of growth hormone gene and growth rate in Indian major carp species.

The phenomenon of growth is a leading factor for aquaculture success. The uneven growth of major Indian carps (Labeo rohita, Catla catla, and Cirrhinus mrigala) is a serious issue in fish culture from an economic point of view. The growth hormone (GH) gene is crucial for selection in commercially cultivated fish species for better growth and production. Indian major carp (L. rohita, C. catla, and C. mrigala) are commonly cultured in Pakistan. The GH expression was examined using qPCR to understand growth in fish species better. Muscle tissue samples (n=480) from 160 individuals of the same age were collected from three species (L. rohita, C. catla, and C. mrigala). Individuals were divided into two groups (high-weight and low-weight groups), cultured under normal conditions. The housekeeping gene ß-actin validated GH expression in fast and slow-growing fishes from the same species. Results showed that GH expression varies across species and fish specimens that overweight their counterpart feature have higher GH expression. A selection for overweight fish in the aquaculture breeding systems is preferable as those fish could inherit their genomics to the future cohort, enhancing production, and commercial profit for farmers. Comprehensive research about different growth genes and the environmental aspects that influence fish growth is mandatory. No work has been reported regarding the growth gene analysis of fish from Pakistan. This report was Pakistan's first and baseline study regarding growth analysis of main culturable fish species at the molecular level.

Impact of different light intensities to Japanese quail eggs on hatching results and post-hatch performance.

The study aims to evaluate dichromatic light with different intensities during incubation on pre- and post-hatch performance of Japanese quail. In completely randomized design, 600 quail hatching eggs were evenly divided into 4 treatments, 3 replicates, and 50 eggs in each. These eggs were provided with lighted incubation [dichromatic light; green-red (GR) with three different intensities 150, 250, and 350 lx] with lighting schedule (12L: 12D), and one treatment was the complete dark period during incubation and considered as control group. After hatch, 300 quail chicks were equally divided into four treatments (3 replicates and 25 chicks in each). Effects of GR light with different intensities during incubation were evaluated on hatching traits and post-hatch performance of Japanese quails. Regarding hatching traits, hatchability was better in all the treated groups (150, 250, and 350 lx of GR light during incubation) compared to complete dark. In terms of embryonic mortality, early embryonic mortality was lower in the 250 lx group, mid embryonic mortality in dark, and late embryonic mortality in the 150 lx group. Furthermore, an ideal chick spread and growth performance (weight gain, feed intake, feed conversion ratio, and liveability) was reported in 250 lx group compared to other treated groups. Serum chemistry values were higher in dark group compared to 150, 250, and 350 lx groups. Pre-slaughter weight was higher in 250 and 350 lx group than 150 lx and dark group, whereas carcass weight was better in 350 lx group compared to dark. In conclusion, stimulation of dichromatic light (green + red) at 250 lx to Japanese quail eggs positively influenced hatching traits and post-hatch growth performance of Japanese quail.

Effects of prenatal dichromatic light exposure on hatching results and post-hatch performance of Japanese quail.

Present study evaluated the effect of lighted incubation on pre- and post-hatch performance of Japanese quail. In a completely randomized design, 1200 eggs were evenly divided into 4 treatments groups having six replicates (each tray was considered as replicate), 50 eggs each. Different dichromatic lights (Green + Red; GR, Green + Blue; GB, and Blue + Red; BR) of 250 lux were provided during incubation for 12 h daily and effects of these lights very evaluated on hatching results and post-hatch growth. After hatch, 600 quail chicks were divided into 4 treatments, 6 replicates, and 25 birds each. Regarding hatching traits, better hatchability was found in the GR group compared to GB, BR, and dark group; while early embryonic mortality was lower in BR, GB, and dark group than GR; mid embryonic mortality was lower in dark group and late embryonic mortality was noted in the GR group than those of other treatment groups. In addition, moisture loss during incubation was minimum in BR and dark groups; however, chick spread was better in the GR group. In terms of growth performance, weight gain was better in the GR group; feed intake in dark, feed conversion ratio in BR, and livability were better in BR and GR group. In morphometrics, keel and shank length were higher in all the colored groups (GB, BR, and GR) whereas body length, wing spread, shank circumference, drumstick length, and circumference were higher in the GR group. Regarding serum chemistry, glucose, albumin, and globulin levels were higher in the GR group. It was concluded that under the experimental conditions, GR light at the prenatal stage to Japanese quail eggs positively influenced hatching performance and post-hatch growth.

Phylogenetic and antigenic analysis of infectious bronchitis virus isolated from commercial and backyard chickens in Pakistan, 2015-2018.

Infectious bronchitis virus (IBV) is a rapidly evolving virus affecting both vaccinated and unvaccinated poultry flocks and is responsible for significant economic losses globally; hence, it is imperative to obtain a deeper understanding of this pathogen. In this study, seven IBV strains were isolated from commercial and backyard poultry flocks during 2015-2018. We obtained full-length IBV genomes of two viruses using the Illumina sequencing method, while five additional viruses were genetically characterized through full-length spike (S1) gene sequencing. Phylogenetic and distance analysis based on complete S1 gene and full-length genome sequences revealed that one IBV isolate belonged to genotype GI-1 and six viruses were clustered within genotype GI-13. Deduced amino acid sequences of GI-13 strains exhibited 31.8-37.2 % divergence with the commonly used classic vaccine strains (M41) and 2.7-12.6 % with variant vaccine strains (4/91) in Pakistan. High evolutionary distances suggest that the IBV viruses circulating in Pakistan are under continuous evolutionary pressure. Moreover, ch/IBV/Pak/AW-2/2017 was found to have originated from an intra-genotypic recombination event between the variant group (GI-23 lineage as a major parent) and variant vaccine strain (4/91-like as a minor parent) and is the first example of recombination within genotype GI-13 in Pakistan. Together, these findings provide genetic and evolutionary insights into the currently circulating IBV genotypes in Pakistan, which could help to better understand the origin, spread and evolution of IBVs, and to ascertain the importance of disease monitoring as well as re-evaluation forof currently used vaccines and vaccination programmes.