Category Archives: b_original_articles

Original Articles

Influence of anaerobic digestate type on nitrogen dynamics and leaching losses across two soils

Sara Bano1, Muhammad Imtiaz Rashid2, Amna Akhtar1, Farhan Hafeez1, Rashid Nazir1, Faridullah1, Muhammad Irshad1, Gabrijel Ondrasek3*, Akhtar Iqbal1*

1Department of Environmental Sciences, COMSATS University Islamabad (CUI), Abbottabad Campus, Tobe Camp, University Road, Abbottabad, Pakistan.

2Center of Excellence in Environmental Studies, King Abdulaziz University, Jeddah 21589, Saudi Arabia.

3Department of Soil Amelioration, Faculty of Agriculture, University of Zagreb, HR-10000 Zagreb, Croatia.


To enhance soil fertility and subsequent crop yields, digestate, byproduct of anaerobic digestion, can serve as a supplement or potential alternative to chemical fertilizers when sensibly utilized. This study assessed the impact of two types of digestates on two distinct soils for the perspective of agriculture (affecting pH, EC, organic carbon and mineral nitrogen) and environment (mineral nitrogen leaching under two rainfall patterns). Both soils mainly differed in silt and sand contents: 32% silt and 47% sand for soil-1 and 42% silt and 39% sand for soil-2. Two sets of controlled experiments served the purpose, in which first set involved a soil incubation experiment, applying two digestates to two soil types at 28°C for 60 days. The second set comprised reconstituted soil columns to collect soil solutions at depths of 2.5 cm and 7.5 cm after rainfall application under two patterns for 45 days. Results indicated that application of both digestates in test soils initially increased pH at day 15, followed by a decrease at days 30 and 60. Both digestates significantly elevated soil electrical conductivity compared to control treatments in both soils. Organic carbon content displayed variable impacts, with a slight decrease for solid digestate (12%) and higher decrease for liquid-amended soil (43%) for soil-1. While a significant decrease was observed for soil-2 throughout the incubation period for both amendments (34% and 36% for solid and liquid amended soils respectively). Rapid nitrification occurred with the application of both digestates in both soils, albeit at different rates. Soil-2 exhibited 1.2 to 2 folds higher net nitrification rate (depending upon digestate type and days of incubation) compared to soil-1. Liquid digestate induced more mineral nitrogen compared to solid digestate in both soils. Interestingly, rainfall frequency, digestate type, and soil type influenced the leaching of ammonium and nitrates, with nitrates recording higher levels in both soils, at both depths, and under both rainfall patterns.

 Keywords: Slurry, Nitrates, Crop productivity, Nitrification rate, Soil nutrition

Lead (Pb) accumulation in rice and its impact on DNA stability

Runglawan Sudmoon1, Arunrat Chaveerach2, Unchaleeporn Ameamsri2, Penkhae Thamsenanupap3,4, Natapol Pumipuntu4,5, Shiou Yih Lee6, Ong Ghim Hock6, Tawatchai Tanee3,4*

1Faculty of Law, Khon Kaen University, Khon Kaen, Thailand

2Department of Biology, Faculty of Science, Khon Kaen University, Khon Kaen, Thailand

3Faculty of Environment and Resource Studies, Mahasarakham University, Maha Sarakham, Thailand

4One Health Research Unit, Mahasarakham University, Maha Sarakham, Thailand

5Faculty of Veterinary Sciences, Mahasarakham University, Maha Sarakham, Thailand

6Faculty of Health and Life Sciences, INTI International University, Negeri Sembilan, Malaysia


Heavy metal contamination, notably lead (Pb), poses a threat to plants and, consequently, human health through the food chain. This study investigates Pb accumulation in rice and how it affects the rice DNA. The rice (Oryza sativa) samples were cultivated in the soil supplemented with Pb at concentrations of 0, 15, 30, 60, and 120 mg/kg. Then the samples were harvested and analyzed for Pb accumulation and DNA alterations using the PCR amplification profiles measured through genomic template stability (GTS). The results demonstrate a Pb concentration hierarchy (root > stem > leaves), with Bioconcentration Factor (BCF) and Translocation Factor (TF) rising when soil Pb contents are supplemented. For DNA alterations, observed GTS values ranged from 23.3 to 76.67%, revealed a general decline with increasing Pb. This correlation assured the influence of Pb on rice DNA stability. Our findings suggest that heavy metal concentration, particularly Pb, has a direct influence on the integrity of rice DNA. Understanding these dynamics is vital for unraveling the complexities of heavy metal-induced genetic changes in plants and their potential implications for food safety and environmental health.

Keywords: Bioconcentration Factor, DNA changes, Genomic template stability, Lead accumulation, Translocation Factor

Bifenthrin induced toxic effects on haematological, reproductive and histo-morphological profile in adult male quail (Coturnix japonica)

Rabia Kalsoom1, Hani Z. Asfour2, Hafiz Muhammad Ali3, Abdul Qayyum3*, Shazia Anjum4**, Faisal Maqbool1, Nuzhat Sial1, Riaz Hussain3, Sultan H. Alamri5, Nadeem Ali6, Nisreen Rajeh7, Irfan Irshad8, Asif Idrees9
1Department of Zoology, The Islamia University of Bahawalpur, 63100, Pakistan
2Department of Medical Microbiology and Parasitology, Faculty of Medicine, King Abdulaziz University, Jeddah-21589, Saudi Arabia
3Faculty of Veterinary and Animal Sciences, The Islamia University of Bahawalpur, 63100, Pakistan
4Faculty of Chemical and Biological Sciences, The Islamia University of Bahawalpur, 63100, Pakistan
5Department of Family Medicine, Faculty of Medicine, King Abdulaziz University, Jeddah-21589, Saudi Arabia
6Center of Excellence in Environmental Studies, King Abdulaziz University, Jeddah-21589, Saudi Arabia
7Department of Clinical Anatomy, Faculty of Medicine, King Abdulaziz University, Jeddah-21589, Saudi Arabia
8Institute of Continuing Education and Extension, University of Veterinary and Animal Sciences, Lahore-54000, Pakistan
9KBCMA, College of Veterinary and Animal Sciences, Narowal, Pakistan


Currently terrestrial ecologies are polluted by numerous chemical compounds unceasingly leading to high risk of exposure to variety of life. Among these, bifenthrin is widely used for the control of different sucking and chewing insects and other leaf miner insects around the globe. The current study elaborated the toxic effects of oral administration of bifenthrin in male Japanese quails at sub-lethal (less than 1% mortality) concentrations (10mg/kg, 20mg/kg and 30mg/kg b.w.) during a period of 30 days. During the course of the study, the quails did not show any behavioural or clinical signs. However, a significantly (p<0.05) decreased RBCs and platelets counts and haemoglobin concentration while a significantly (p<0.05) increased MCV, MCHC, total leucocytes and neutrophils were observed in the birds administered with higher concentrations of bifenthrin. Moreover, the incidence of lobed nuclei, blebbed nuclei, condensed nuclei, notched nuclei, binuclear, pear shaped and micro-nuclei were significantly (p<0.05) increased in the erythrocytes of the groups C and D during the experiment. The diameter of seminiferous tubules, height of germinal epithelium and the number of seminiferous tubules containing normal spermatozoa were significantly (p<0.01) decreased while the number of pyknotic cells and degenerated seminiferous tubules were increased significantly (p<0.05) towards the end of the experiment (day-30) in the quails of groups C and D compared to the control group. Hence, the alterations in the hematological indices and histopathological changes in heart, spleen and testes indicate potential toxicity of bifenthrin and its adverse effects in the Japanese quail even at sub-lethal concentrations.

 Keywords: Bifenthrin, Blood profile, Nuclear alterations, Seminiferous tubules, Histo-pathology, Testes

The differential regulation of tumor suppressor genes (SAMD9, SPRED1, TGFBI, DUSP6, CDX2, TP53) and MAPK/ERK signaling pathway in colorectal cancer

Mahmood Rasool1*, Khalid I Alhassan2, Sajjad Karim1, Absarul Haque3, Mohammed HZ Mutwakil2, Mohammed Alharthi4, Adeel G Chaudhary1, Peter Natesan Pushparaj1

1Center of Excellence in Genomic Medicine Research, Faculty of Applied Medical Sciences, King Abdulaziz University, Jeddah, Saudi Arabia

2Department of Biological Science, King Abdulaziz University, Jeddah, Saudi Arabia

3King Fahd Medical Research Center, Faculty of Applied Medical Sciences, King Abdulaziz University, Jeddah, Saudi Arabia

4Faculty of Medicine, King Abdulaziz University Hospital, King Abdulaziz University, Jeddah, Saudi Arabia


Despite considerable advancements in research, particularly in oncology, colorectal cancer (CRC) remains a formidable and deadly disease. It is crucial to delve deeper into the effects of targeted therapies, signaling pathways, and genetic regulation to ensure the effectiveness of cancer treatments. In this study, we obtained microarray data for four patient-derived CRC organoids from the Gene Expression Omnibus (GEO) database (accession number: GSE114060). We then used several next-generation knowledge discovery (NGKD) tools, such as GEO2R, Metascape, WebGestalt, and Ingenuity Pathway Analysis (IPA) software, to investigate the underlying molecular mechanisms in colorectal cancer-derived organoids treated with trametinib compared to those treated with DMSO. Our NGKD analysis revealed upregulation of SAMD, TP53, and SPTLC3 and downregulation of SPRED1, TGFBI, and DUSP6. The MAPK/ERK signaling pathway was significantly downregulated and was associated with reduced expression of CDX2 in CRC organoids treated with trametinib. We concluded that SAMD9, SPRED1, TGFBI, TP53, and DUSP6 are differentially regulated and can play a pivotal role in the downregulation of the MAPK/ERK signaling pathway to suppress CRC. The use of targeted therapies to regulate the specific gene signatures identified in the current study may be beneficial in the CRC associated tumor suppression . On the other hand, upregulation of SPTLC3 may be induced by MEK inhibitors and may cause hepatotoxicity alongside nonalcoholic steatohepatitis.

Keywords: Colorectal cancer, Organoids, Tumor suppression, MAPK/ERK signaling pathway, MEK inhibitor, Hepatotoxicity

The roles of FGFR, EGFR and AMP-activated protein kinase pathway in colorectal cancer stem cells derived spheroids: Implications in colorectal cancer treatment

Mahmood Rasool1*, Khalid I. Alhassan2, Sajjad Karim1, Absarul Haque3, Mohammed H.Z. Mutwakil2, Mohammed Alharthi4, Adeel G. Chaudhary1, Peter Natesan Pushparaj1

1Center of Excellence in Genomic Medicine Research, Faculty of Applied Medical Sciences, King Abdulaziz University, Jeddah, Saudi Arabia

2Department of Biological Science, King Abdulaziz University, Jeddah, Saudi Arabia

3King Fahd Medical Research Center, Faculty of Applied Medical Sciences, King Abdulaziz University, Jeddah, Saudi Arabia

4Faculty of Medicine, King Abdulaziz University Hospital, King Abdulaziz University, Jeddah, Saudi Arabia


The aim of our research was to identify molecular targets that can be targeted by drugs and patient-specific models for personalized medicine for colorectal cancer (CRC). Here, we obtained high-throughput RNA sequencing data from Gene Expression Omnibus (GEO) with accession number GSE205787 and analyzed it using next-generation knowledge discovery tools such as BioJupies and Ingenuity Pathway Analysis (IPA) software. Differentially expressed genes (DEGs) were identified by comparing the raw counts from 47 CRC patient-derived spheroids (CRC-CSCs) with those from normal spheroids from the epithelium of the colon and rectum of healthy individuals, using BioJupies tools. IPA was used to identify differentially regulated canonical pathways, upstream regulators of CRC, non-directional networks, diseases, and biofunctions, as well as to conduct subsequent perturbation analysis using the Molecular Prediction Analysis (MAP) tool. Our study demonstrates that several KEGG pathways, including the AMPK, Phospholipase D, MAPK, and PI3-AKT signaling pathways, were significantly downregulated in the CRC-CSC group. Additionally, Wnt signaling and FGFR pathways were significantly upregulated. Moreover, according to Wikipathways, the EGF/EGFR signaling pathway, MAPK signaling pathway, G-protein signaling pathway, and Focal Adhesion-PI3-AKT pathway were downregulated in the CRC-CSC group. Furthermore, based on the Reactome, the Metabolism, Vesicle-mediated transport, RAF signaling, and G-alpha (12/13) signaling pathways were also downregulated in the CRC-CSC group. Utilizing innovative drug combination approaches and innovative drug delivery techniques, CRC treatments can be enhanced by modulating the FGFR, EGFR, and AMPK signaling pathways, which may ultimately lead to improved patient outcomes.

Keywords: Colorectal cancer, Cancer stem cell spheroids, AMP‐activated protein kinase signaling, Fibroblast Growth Factor Receptor, Epidermal Growth Factor Receptor, BioJupies, Ingenuity Pathway Analysis

Antibacterial and lignocellulose-degrading enzyme activities of coprophilous fungi obtained from cow dung in Thailand

Narumon Tangthirasunun1*, Darbhe Jayarama Bhat2,3 , Supattra Poeaim1

1Department of Biology, School of Science, King Mongkut’s Institute of Technology Ladkrabang, Bangkok, 10520, Thailand

2Department of Botany and Microbiology, College of Science, King Saud University, P.O. Box 2455, Riyadh, 11451, Saudi Arabia

3Vishnugupta Vishwavidyapeetam, Ashoke, Gokarna, 581326, India


Twenty-seven coprophilous fungi, isolated from field-fed cow dung in an organic farm in Thailand, were identified using morphology and ITS barcode. A total of five genera viz. Aspergillus, Hamigera, Paecilomyces, Penicillium, and Talaromyces were identified with varying numbers and growth rates. These fungi were evaluated for their antibacterial properties against Gram-positive (Bacillus subtilis, Kocuria rhizophila, Staphylococcus aureus and St. epidermidis) and Gram-negative (Escherichia coli and Pseudomonas aeruginosa) bacteria. Pe. javanicum NTD-SP2-01 and Talaromyces sp. NTD-SP5-48 exhibited activity against all bacteria when tested with agar plug diffusion method. Talaromyces sp. NTD-SP5-48 was particularly effective against Gram-negative bacteria. As. terreus NTD-NG1-05 displayed the highest activity against five bacterial strains, except Ps. aeruginosa. Notably, As. terreus NTD-NG1-05 and Talaromyces sp. NTD-SP5-48 demonstrated extended antibacterial activity in the agar disk diffusion method, with fermented broth (FB) showing superior inhibitory effects compared to mycelial extract (MY). Both isolates demonstrated significant antibacterial activity against B. Subtilis. Furthermore, all isolates exhibited significant antibacterial activity against B. subtilis, with a diffusion of 0.125 mg/disk. Only Talaromyces sp. NTD-SP5-48 (FB) displayed the highest inhibition activity against Ps. aeruginosa, with a diffusion of 1 mg/disk (100 mg/mL). In terms of enzyme activity, all isolates exhibited cellulase activity, with Talaromyces sp. showing the highest cellulase activity, followed by As. terreus. Laccase activity was only observed in the unidentified isolate NTD-SP5-34, while none of the isolates showed pectinase activity.

Keywords: Antimicrobial activity, Coprophilous fungi, Heat-resistant fungi, Lignocellulolytic enzymes

Comparative study of productive and reproductive parameters of Holstein Friesian cows in different agroecological zones under subtropical conditions of Pakistan

Masood Ur Rehman1, Syed Muhammad Suhail1, Iqbal Munir2, Rajwali Khan1*

1Department of Livestock Management, Breeding and Genetics, The University of Agriculture, Peshawar, 25130, Khyber Pakhtunkhwa, Pakistan

2Institute of Biotechnology and Genetic Engineering, The University of Agriculture, Peshawar, 25130, Khyber Pakhtunkhwa, Pakistan


The current study aimed to evaluate the impact of three different agroecological zones on the productive and reproductive performance of Holstein-Friesian (HF) cows, along with their comparative analysis of physiological and molecular markers under subtropical conditions of Pakistan. The productive (milk yield and composition) and reproductive (service per conception and calving interval) performance of genetically identical (n=210) HF cows placed across three agroecological zones: irrigated lowlands (Okara), wet (Abbottabad), and dry (Queta) highlands were analyzed. Additionally, heat stress markers (superoxide dismutase, glutathione peroxidase, serum cortisol, bovine heat shock protein 70 (HSP-70), and blood glucose) from five cows in each location (n=15 in total) were also investigated in early and late summers using commercial ELISA and calorimetric kits. Furthermore, the mRNA levels of heat shock protein genes HSPA8 and HSP90AB1 were also quantified in three agroecological zones through qRT-PCR.  The results revealed that cows raised in the wet highlands of Abbottabad exhibited significantly higher (P<0.05) milk production (daily and total lactational yield), and the lowest service period, calving interval and number of services per conception as compared to the cows from the other two agroecological zones. Furthermore, there was a significant effect (P<0.05) of agroecological zones on the heat stress indicators (SOD, GPX, cortisol, blood glucose, and HSP70) profiles and expression of heat shock protein genes in HF cows. The dairy cows from Abbottabad (Wet highlands) showed a significantly lower profile of heat stress indicators as compared to the cows from the other two agroecological zones. It can be inferred that HF cows function better in moderate agroecological zones of subtropical countries.

Keywords: Heat stress indicators, Subtropics, Holstein-Friesian cows, Agroecological zone

Moringa leaf extract enhances the growth and yield characteristics of buckwheat genotypes by modulating the biochemical and physiological activities

Heer Baloch1, Irfan Ali Sabir2, Saadullah Khan Leghari1, Muhammad Sohail Saddiq3*, Pravej Alam4, Shahbaz Khan5*, Esha Mehik Fatima6, Mateen Sajid7, Muhammad Hammad Raza8, Muhammad Arif Hussain1, Muhammad Ayoub1, Rashid Iqbal9

1Department of Botany, Ghazi University, Dera Ghazi Khan, Pakistan

2College of Horticulture, South China Agricultural University, Guangzhou, 510642, China

3Department of Agronomy, Ghazi University, Dera Ghazi Khan, Pakistan

4Department of Biology, College of Sciences and Humanities, Prince Sattam Bin Abdulaziz University, Al- Kharj 11942, Saudi Arabia

5Colorado Water Center, Colorado State University, Fort Collins, CO, USA

6Department of Entomology, Muhammad Nawaz Shareef University of Agriculture, Multan, Pakistan

7Department of Horticulture, Ghazi University, Dera Ghazi Khan, Pakistan

8Department of Agri. Extension and Education, Ghazi University, Dera Ghazi Khan, Pakistan

9Department of Agronomy, Faculty of Agriculture and Environment, The Islamia University of Bahawalpur, Pakistan


Moringa leaf extract (MLE) as a biostimulant has demonstrated success in boosting the productivity of several agronomic crops, but its impact on Buckwheat crops remains unexplored. Buckwheat, recognized as an essential nutritional and functional food crop, often exhibits lower yields compared to major cereal crops grown in similar environments. Therefore, this research aimed to investigate the impact of different concentrations (1%, 2%, and 3%) of MLE on the agricultural performance of common buckwheat (CB) and tartary buckwheat (TB). A pot experiment was carried out according to completely randomized design with factorial arrangements having three replications. Results demonstrated significant improvements in growth parameters (branches, leaves, nodes, and internodes) for MLE-treated plants compared to the control group. Foliar treatment MLE 2% also increased chlorophyll content, improved membrane stability index (MSI) and relative water content (RWC), and enhanced biochemical composition (phenolic compounds, free amino acids, leaf proline, and soluble sugars) in both buckwheat genotypes followed by MLE 3% and MLE 1%. TB produced significantly higher grain yield (0.74 g) as compared to CB (0.43 g). The findings showed that a foliar treatment of MLE 2% led to increased grain yield in both TB (0.97 g) and CB (0.55 g) as compared to control group plants (0.37g TB and 0.22 g CB) respectively. This increase was associated with elevated activities of photosynthetic pigments, phenolic content, RWC, free amino acids, soluble sugars, and catalase in both buckwheat genotypes. In conclusion, MLE application at 2% significantly boosted the agriculture performance of buckwheat, and this study unlocked new insights into optimizing the productivity of the vital food crop.

Keywords: Biostimulant, Free amino acid, Phenolic, Buckwheat, Foliar application, Grain yield

Development of DNA vaccine(s) against Mycobacterium specific genes and prime boost with BCG

Aeman Jilani1, Mirza Imran Shahzad1*, Muhammad Mohsin Zaman1, Areeba Yousaf1, Gildardo Rivera2

1Institute of Biochemistry, Biotechnology and Bioinformatics, The Islamia University of Bahawalpur, Pakistan

2Laboratorio de Biotecnología Farmacéutica, Centro de Biotecnología Genómica, Instituto Politécnico Nacional, México


Tuberculosis (TB), the white plague of Europe is still uncontrolled and fatal in many parts of the world including Pakistan. It is a major cause of morbidity and mortality in human and domestic animals in Pakistan. No new vaccine in the last hundred years has been developed except for a few encouraging results from recombinant and DNA vaccines in the past two decades. Five Mycobacterium specific genes (Rv0379, Rv3914, Rv3006, Rv0432+SP, and Rv0432-SP) were selected to develop DNA vaccine(s). All the constructs were tested on mice using both naked DNA and prime-boost methodologies. Forty-five BALB/c mice were divided into three main groups; DNA vaccine group, BCG Prime boost group, and Control group. Post-vaccine (PV) and post-challenge (PC) immune responses were confirmed through cytokine ELISA and qRT PCR. IFN-γ was additionally checked in plasma as well. Based on cytokine ELISA PC immune responses showed significant differences in TNF-α levels for both naked DNA vaccine groups (Rv0379, Rv3006, and Rv0432-SP) and BCG primed Rv3914 group in comparison to the BCG control group (p<0.05). Based on qRT PCR, IL-6, TNF-α, IFN-γ, and IL-1β showed no significant difference among all the vaccines and BCG control groups (CT range 25- 30). IFN-γ levels in plasma were analyzed PC; two vaccines Rv3006/LppZ and BCG primed Rv0432/SodC-SP (highest mean value 1360.35 pg/ml) have shown significant results (cutoff value 21pg/ml) at 63 days. All the vaccine construct(s) alone or in combination have significant therapeutic effects in comparison to BCG and negative control groups.


Keywords: Mycobacterium tuberculosis, DNA vaccine, Tumor necrosis factor-alpha, Interleukin-6, Interferon-gamma, Interleukin-1beta, Bacille Calmette-Guérin (BCG)

Hemato-biochemical changes, molecular characterization and phylogenetic analysis of the 2022 Lumpy Skin Disease (LSD) outbreak in Cholistan, Pakistan

Tayyaba Asghar1, Mudassar Mohiuddin1*, Ayesha Mohiud Din2,3, Tahira Kamal4, Muhammad Khalid Mansoor1,

Abubakar Siddique5,6,7, Mudasser Habib8, Riaz Hussain9, Muhammad Taslim Ghori10, Hira Hameed1, Humaira Rizwana11,

Islem Abid12, Muhammad Ehsan13, Aroosh Shabbir14

1Department of Microbiology, Faculty of Veterinary and Animal Sciences, The Islamia University of Bahawalpur, Pakistan

2Department of Biotechnology, Virtual University of Pakistan, Lahore, Pakistan

3School of Biological Sciences, University of Southampton, Southampton, UK

4National Institute for Genomics and Advanced Biotechnology, National Agricultural Research Centre (NARC), Islamabad, Pakistan

5Atta-ur-Rahman School of Applied Biosciences (ASAB), National University of Science and Technology (NUST), Islamabad, Pakistan

6Department of Veterinary Medicine, College of Animal Sciences, Zhejiang University, China

7Hainan Institute of Zhejiang University, Sanya, China

8Animal Sciences Division, Nuclear Institute for Agriculture and Biology, Faisalabad, Pakistan

9Department of Pathology, Faculty of Veterinary and Animal Sciences, The Islamia University of Bahawalpur, Pakistan

10Department of Clinical Medicine and Surgery, Faculty of Veterinary and Animal Sciences, The Islamia University of Bahawalpur, Pakistan

11Department of Botany and Microbiology, College of Science, King Saud University, Riyadh 11451, Saudi Arabia

12Centre of Excellence in Biotechnology Research, King Saud University, P.O Box 2455, Riyad 11495, Saudi Arabia

13Department of Parasitology, Faculty of Veterinary and Animal Sciences, The Islamia University of Bahawalpur, Pakistan

14Institute of Molecular Biology and Biotechnology (IMBB), The University of Lahore, Pakistan


The lumpy skin disease virus (LSDV) is a member of the Capripox genus of the Poxviridae family. It is the causative agent of lumpy skin, a highly contagious disease of cattle, water buffalo, sheep, and goats. In 2022, several outbreaks of LSD were reported in the Cholistan region of Pakistan, which has a large population of livestock living in arid conditions. A total of 230 blood/serum and scab samples were collected from three LSD outbreak locations. Clinically, affected cattle showed acute clinical signs characterized by skin nodules, fever, enlarged lymph nodes, emaciation, and lower leg edema. Hematological findings revealed non-significant changes in red blood cell and white blood cell counts (some animals had leukocytosis while others were leukopenic) whereas, hemoglobin level were significantly low. Platelet count, MPV, PCT, P-LRC, and P-LCC were elevated. Granulocytes were significantly low in LSD affected cattle while lymphocyte counts were significantly high. Serological findings revealed elevated protein levels, along with high creatinine and ALT concentrations. Amplification of DNA-dependent RNA polymerase 30 kDa subunit gene (RPO30) confirmed the presence of LSD virus in all suspected samples. Phylogenetic analysis showed that all Pakistani isolates clustered closely with isolates from neighboring countries. The SNPs differences were less than 20 among these isolates, indicating their close resemblance with each other. It can, therefore, be inferred that our LSD strains might be originated from neighboring Asian countries, that were affected by LSD in previous years.

Keywords: Lumpy skin disease virus (LSDV), Cholistan, Skin lesions, Hematology, Phylogenetic analysis