Validation and application of a method for determination of multi-class pesticides in muscle chicken breast fillets using QuEChERS extraction and GC/MS.

Introduction: The occurrence of pesticide residues in animal products deserves attention because of the contamination by environmental pollutants and pesticides that may be present in the food that animals are fed. The goal of this work was the validation of a method for detection of residues of multiple classes of pesticide and determination of their residues in chicken breast fillets. Material and Methods: Gas chromatography with mass spectrometry was used for analysis. A modified quick, easy, cheap, effective, rugged and safe (QuEChERS) method was put into practice for its validation and applied to real samples. The study optimised mass detection and investigated the effect of a freezing step during the preparation of samples. Pesticides were determined in samples from conventional and organic production. Results: The impact of the matrix effect decreased, with the largest number of pesticides and satisfactory recovery determined by the application of mixed solvent acetonitrile and ethyl acetate for extraction. Detection of pesticide residues was achieved in a linear range between 5 and 50 µg/kg with satisfactory excellent correlation coefficients greater than 0.99. The recovery of all the pesticide residues ranged between 71.2 and 118.80%. The relative standard deviation was from 2.9% to 18.1% for all validated pesticide residues. The limits of quantification were in the range of 3.0-4.9 µg/kg. Out of 56 pesticide residues analysed in real samples, 5 were detected: α endosulfan, cypermethrin, endosulfan sulphate, permethrin and p,p´-dichlorodiphenyltrichloroethane (DDT) and their concentrations ranged from 4.9 to 15.2 µg/kg. Conclusion: All tested samples were compliant with the evaluation criteria, and detected values of pesticide residues were lower than the maximum residual levels.

Molecular Detection of Porcine Cytomegalovirus, Porcine Parvovirus, Aujeszky Disease Virus and Porcine Reproductive and Respiratory Syndrome Virus in Wild Boars Hunted in Serbia during 2023.

Porcine cytomegalovirus (PCMV) infection is widespread worldwide and has a high prevalence in swine herds, especially in countries with intensive swine production. PCMV is zoonotic and can impact xenotransplants. It is the third swine virus known to be zoonotic, following swine influenza virus (influenza A) and hepatitis E virus genotype 3 (HEVgt3 or HEV-3). Wild boars, serving as reservoirs for various pathogens, including PCMV, pose a risk to both the pig industry and public health. This study aimed to investigate PCMV infection in Serbian wild boars using real-time PCR and assess other viral infections. We also tested samples for the presence of other viral infections: Aujeszky disease virus (ADV), Porcine parvovirus (PPV) and Porcine reproductive respiratory syndrome (PRRSV). Samples from 50 wild boars across 3 districts were tested. Results showed 8% positivity for PCMV DNA, with females showing higher infection rates. Porcine parvovirus (PPV) was detected in 56% of samples, while Porcine reproductive respiratory syndrome virus (PRRSV) was absent. ADV was found in 18% of samples, primarily in younger animals. This research contributes to understanding PCMV prevalence in Serbian wild boars and emphasizes the importance of monitoring viral infections in wild populations, considering the potential zoonotic and economic implications.

Bovine Parainfluenza Virus 3 and Bovine Respiratory Syncytial Virus: Dominant Viral Players in Bovine Respiratory Disease Complex among Serbian Cattle.

Bovine respiratory disease complex, a complex respiratory ailment in cattle, results from a combination of viral and bacterial factors, compounded by environmental stressors such as overcrowding, transportation, and adverse weather conditions. Its impact extends beyond mere health concerns, posing significant economic threats to the cattle industry. This study presents an extensive investigation into viral pathogens associated with BRDC in Serbian cattle, utilizing serum samples and nasal swabs. A cross-sectional study was conducted in 2024 across 65 randomly selected dairy farms in Serbia, excluding farms with vaccinated cattle. The farms were categorized by their livestock count: small (≤50 animals), medium (51-200 animals), and large (>200 animals). Serum samples from adult cattle older than 24 months were tested for antibodies against BVDV, BHV-1, BRSV, and BPIV3. Nasal swab samples from the animals with respiratory signs were tested using PCR for viral genome detection. The results showed seropositivity for all four viruses across all of the farms, with BPIV3 exhibiting universal seropositivity. Medium-sized and large farms demonstrated higher levels of seropositivity for BRSV and BHV-1 compared to small farms (p < 0.05). Our true seroprevalence estimates at the animal level were 84.29% for BRSV, 54.08% for BVDV, 90.61% for BHV-1, and 84.59% for BPIV3. A PCR analysis of the nasal swabs revealed positive detections for BRSV (20%), BHV-1 (1.7%), BVDV (8%), and BPIV3 (10.9%). Influenza D virus was not found in any of the samples. This study provides critical insights into the prevalence and circulation of viral pathogens associated with BRDC in Serbian cattle, emphasizing the importance of surveillance and control measures to mitigate the impact of respiratory diseases in cattle populations.

Presence of Mycoplasma bovis in Bulk Tank Milk and Associated Risk Factor Analysis in Serbian Dairy Farms.

Mycoplasma bovis (M. bovis) is a significant pathogen responsible for highly transmissible mastitis in cattle globally. It primarily spreads through colostrum, milk, and semen. Cows with persistent infections act as carriers, intermittently releasing the pathogen, making their milk a pivotal factor in infection transmission. Given the limited seroprevalence surveys in Serbia, this study aimed to detect M. bovis presence in bulk tank milk (BTM), determine route shedding, and evaluate infection risks. BTM samples were collected from 115 dairy farms across Serbia, with M. bovis DNA detected in 11 out of the 115 samples by real-time PCR. Additionally, M. bovis was detected in 1.30% of nasal swabs sampled from apparently healthy animals. A univariate analysis of the risk factors associated with M. bovis presence in the BTM samples revealed correlations with factors such as the breed, farm seropositivity, pre-milking and post-milking disinfection practices, farm type, cow population, milk yield, number of cows in the BTM samples, and parity. Seropositive farms exhibited the highest likelihood of M. bovis presence in milk. Moreover, pre- and post-milking disinfection practices and highly productive cows yielding over 8000 L of milk were identified as risk factors for PCR-positive BTM. In a multivariable mixed regression analysis, a risk factor for the presence of M. bovis infection in the BTM sample was the Holstein breed. These findings underscore a relatively high prevalence of M. bovis in BTM within Serbian dairy farms, suggesting a potential risk for M. bovis spreading through milk and oral route of calves' infection.

Molecular characterization of Canine parvovirus type 2 from diarrheic dogs in Serbia from 2008 to 2020.

Canine parvovirus 2 (CPV-2) is the causal agent of canine parvovirosis an infectious disease with the high fatality rate among dogs. However, in Serbia, it has never been investigated thoroughly. This study was conducted on samples collected from dogs with diarrhea in anamnesis, submitted for various reasons to the Institute of Veterinary Medicine of Serbia, and stored in the sample bank. In total, 50 rectal swab samples were collected from the period 2008 to 2020, and consequently tested. Out of 50 rectal swab samples, the CPV-2 genome was detected in 14 (28%). This retrospective study showed the presence of three different subtypes of CPV-2 in diarrheic dogs during the last 12 years in Serbia. CPV-2a was the most prevalent subtype (60%) followed by CPV-2b (30%), and CPV-2c (10%). Interestingly, CPV-2a had been the predominantly detected subtype up until 2018. Nevertheless in 2019, there was the first detected occurrence of the CPV-2b, followed by the first detection of the CPV-2c in 2020. This study reports the evidence and distribution of CPV-2 from 2008 to 2020, providing new information about the presence of virus strains in Serbia.