S evere fever with thrombocytopenia syndrome (SFTS) is an emerging tickborne disease caused by the SFTS virus (SFTSV; genus Banyangvirus, family Phenuiviridae, order Bunyavirales). The disease is prevalent in East Asia countries. It was first detected in China in 2009 and later in Japan and South Korea (1) and is suspected to be widely spread across other parts of the world (2). The recent identification of SFTSV in Xinjiang, China (3), expanded our awareness of epidemic areas of SFTS and suggested the possibility of SFTSV spreading to bordering countries like Pakistan. However, the presence of SFTSV in Pakistan has been unclear. We investigated the seroprevalence of SFTSV in humans in Pakistan. The Study For this study, we randomly collected human serum samples (n = 1,657) from 4 provinces in Pakistan during 2016-2017 (Figure). All participants were farmers of livestock (sheep, goats, cattle, buffaloes, and camels). We recorded and summarized testing results by sex, age, and geographic location (Table). The collection of human serum samples and subsequent tests were reviewed and approved by the Ethics Committees of

Virulent viruses of the panzootic Avian avulavirus 1 (AAvV-1) of sub-genotype VIIi were repeatedly isolated (2011-2016) from commercial chickens and from multiple non-poultry avian species in Pakistan. These findings provide evidence for the existence of epidemiological links between Newcastle disease outbreaks in commercial poultry and infections with virulent AAvV-1 strains in other avian species kept in proximity to poultry. Our results suggest that the endemicity of Newcastle disease in Pakistan involves multiple hosts and environments.

Middle East Respiratory Syndrome Coronavirus (MERS-CoV) is a zoonotic pathogen capable of causing severe respiratory disease in humans. Although dromedary camels are considered as a major reservoir host, the MERS-CoV infection dynamics in camels are not fully understood. Through surveillance in Pakistan, nasal (n = 776) and serum (n = 1050) samples were collected from camels between November 2015 and February 2018. Samples were collected from animal markets, free-roaming herds and abattoirs. An in-house ELISA was developed to detect IgG against MERS-CoV. A total of 794 camels were found seropositive for MERS-CoV. Prevalence increased with the age and the highest seroprevalence was recorded in camels aged [ 10 years (81.37%) followed by those aged 3.1-10 years (78.65%) and B 3 years (58.19%). Higher prevalence was observed in female (78.13%) as compared to male (70.70%). Of the camel nasal swabs, 22 were found to be positive by RT-qPCR though with high Ct values. Moreover, 2,409 human serum samples were also collected from four provinces of Pakistan during 2016-2017. Among the sampled population, 840 humans were camel herders. Although we found a high rate of MERS-CoV antibody positive dromedaries (75.62%) in Pakistan, no neutralizing antibodies were detected in humans with and without contact to camels.

Bovine brucellosis remains a persistent infection in ruminants in Pakistan. A total of 828 (409 buffaloes and 419 cattle) sera were collected from 11 institutional-owned livestock farms in Punjab, Pakistan. The samples were tested by rose bengal plate agglutination test (RBPT) and indirect enzyme-linked immunosorbent assay (iELISA). The seroprevalence along with 95% confidence interval (CI) was determined. Univariable and multivariable analysis of the epidemiological background data was conducted and odds ratio (OR) was calculated to understand any association between the risk factors and the seroprevalence. An overall seroprevalence of 3.9% (Positive/Tested = 32/828) and 3.3% (27/828) was detected by RBPT and iELISA, respectively. The seroprevalence of 5.6% (CI 3.6–8.3) and 4.7%, (CI 2.8–7.2) and the odds ratio of 2.63 (CI 1.20–5.77) and 2.50 (CI 1.08–5.78) for testing positive by RBPT and iELISA, respectively were significantly higher (p < 0.05) in buffaloes than in cattle. Breed, sex, history of abortion and retention of fetal membranes (RFM) in the animals were not found statistically significantly associated with the infection. RBPT and iELISA based results agreed almost perfect (k = 0.877). In total, Brucella abortus-DNA (9/27) was amplified from seropositive samples by real-time polymerase chain reaction. This study identified for the first time the etiological agents of brucellosis at a molecular level at institutional-owned livestock farms in Pakistan.

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Mycoplasma mastitis is often difficult to control due to a lack of rapid and accurate diagnostic tools. The aim of the current study was to develop a loop-mediated isothermal amplification (LAMP) assay for the detection of Mycoplasma bovis (M. bovis) in mastitic milk. The assay was developed using primers designed for three different target genes: uvrC, 16S rRNA, and gyrB, and validated using mastitic milk samples previously found positive for the target pathogen. Specificity of the developed assay was determined by testing cross-reactivity of LAMP primers against closely related bovine mastitis bacterial pathogens. The sensitivity was found to be higher compared to conventional polymerase chain reaction (PCR). The LAMP assay was also capable of detecting M. bovis in PCR-negative milk samples of cows with clinical mastitis. The uvrC primers were found to be more sensitive, while gyrB primers were more specific; however, 16S rRNA primers were less specific and sensitive compared to either uvrC or gyrB primers. Cohen's kappa values for uvrC, gyrB, and 16S rRNA primers used in the LAMP assays were 0.940, 0.970, and 0.807, respectively. There was a high level of agreement between the test results and the true-disease status as indicated by the receiver operating characteristic (ROC) curve. Our findings suggest that the newly developed LAMP assays targeting the uvrC and gyrB genes could be a useful tool for rapid and accurate diagnosis of mastitis caused by M. bovis.

Omega 3 and 6 fatty acids were enriched in the olein fraction of chia oil via dry fractionation at -30°C. Concentrations of C18:3 (linolenic acid, ω-3) and C18:2 (linoleic acid, ω-6) were 78.16 and 25.42% in the olein fraction. HPLC characterization indicated enrichment of caffeic and chlorogenic acids, quercetin, and phenolic glycosides in the olein fraction. Total antioxidant activities of chia oil and the olein and stearin fractions were 42.5, 53.8, and 34.6%, respectively. After 6 months of storage at 4°C, the concentration of ω-3 in the olein fraction decreased from 78.19 to 76.16%, with a 10% decrease in the ω-3 concentration when the olein fraction was stored at 25°C. The stearin fraction of chia oil exhibited the longest induction period, followed by chia oil and the olein fraction. Amounts of ω-3 and 6 fatty acids can be enriched in the olein fraction from 11.92 and 61.28% to 15.22 and 72.16%, respectively, with reasonable storage stability at low temperature.

Effect of supplementing cheddar cheese with chia oil on omega fatty acids, phenolic compounds, and lipolysis of cheddar cheese was investigated. Milk fat was partially replaced with chia oil, i.e., 2.5, 5, 7.5, and 10% (T1, T2, T3, and T4). Cheese prepared from 100% milk fat served as control, ripened at 6 °C for 90 days. Concentration of α‐linolenic acid in control and T3 was 0.51 and 12.55%. HPLC characterization revealed the concentrations of chlorogenic acid, caffeic acid, quercetin, phenolic glycoside‐k, and phenolic glycoside‐Q in T3 were 0.15, 0.26, 0.62, 1.55, and 1.97 mg/mL. Concentration of cholesterol in 90 days ripened control and T3 was 119 and 92 mg/100 g with lower concentration of organic acids and no difference in sensory characteristics of cheddar cheese up to T3 level. These results suggest that concentration of omega fatty acids and phenolic compounds can be enhanced in cheddar cheese by supplementing with chia oil. Practical applications Health benefits associated with the intake of omega fatty acids and natural antioxidants are scientifically established, demand for functional foods is increasing throughout the world. Cheddar cheese can be supplemented with chia oil to enhance the concentration of beneficial omega fatty acids and phenolic compounds.