Appraisal and validation of a method used for detecting heavy metals in poultry feed in Bangladesh

Research (Published online: 17-09-2022)

13. Appraisal and validation of a method used for detecting heavy metals in poultry feed in Bangladesh

Md. Mosharaf Hossain, Abu Sayeed Md. Abdul Hannan, Md. Mostofa Kamal, Mohammad Abul Hossain, and Shamshad B. Quraishi

Veterinary World, 15(9): 2217-2223

ABSTRACT

Background and Aim: Low concentrations of heavy metals are toxic and pose a serious threat to human health and the environment. Therefore, regular assessments of the toxic metal content in poultry feed are crucial for evaluating feed quality and customer safety. It is difficult to determine the heavy metals in the poultry feed at the trace amount. Therefore, this study aimed to validate this method through the detection of three heavy metals, chromium (Cr), cadmium (Cd), and lead (Pb), in poultry feed samples.

Materials and Methods: Graphite furnace atomic absorption spectrometry (GF-AAS) method was used to analyze the heavy metals in poultry feed according to the guidelines given by the Council Directive 333/2007/EC, Commission Decision 657/2002/EC. In this study, various parameters such as linearity check, limit of detection (LOD), limit of quantification (LOQ), recovery percentage, precision checks, repeatability, reproducibility, and uncertainty measurement were considered to validate and assess the method following international guidelines. Heavy metals, such as Pb, Cr, and Cd, were analyzed from the feed samples in the laboratory using the GF-AAS method (Model: AA-7000 Shimadzu, Japan) with high purity argon as the inert gas, and the absorbance was read at wavelengths of 283.0, 357.9, and 228.8 nm, respectively.

Results: The coefficient of variation (CV%) for system suitability and precision data was <10% for all the metals (Pb, Cr, and Cd) detected in this study. The overall CV% of repeatability and reproducibility ranged from 8.70% to 8.76% and 8.65% to 9.96%, respectively. The linearity of the calibration curves was excellent (r2 > 0.999) at various concentration levels for the three different metals. The recovery (%) was found to be 94.53, 93.97, and 101.63% for Pb, Cr, and Cd, respectively. The LOD values in feed were 0.065, 0.01, and 0.11 mg/kg, and the LOQ values were 0.22, 0.03, and 0.38 mg/kg for Cr, Cd, and Pb, respectively. The values recorded for the measurement uncertainty (%) were 11.48, 4.43, and 12.42% for Cr, Cd, and Pb, respectively.

Conclusion: The results show that these study criteria or parameters have met the validated or acceptable range. Therefore, it is a reliable technique that can be used undoubtedly for the routine analysis of heavy metals in poultry feed samples across the globe.

Keywords: graphite furnace atomic absorption spectrometry, method validation, poultry feed, toxic metals (Pb, Cr, and Cd).

Multiple gene editing in porcine embryos using a combination of microinjection, electroporation, and transfection methods

Research (Published online: 16-09-2022)

12. Multiple gene editing in porcine embryos using a combination of microinjection, electroporation, and transfection methods

Quynh Anh Le, Manita Wittayarat, Zhao Namula, Qingyi Lin, Koki Takebayashi, Maki Hirata, Fuminori Tanihara, Lanh Thi Kim Do, and Takeshige Otoi

Veterinary World, 15(9): 2210-2216

ABSTRACT

Background and Aim: Mosaicism – the presence of both wild-type and mutant alleles – is a serious problem for zygotic gene modification through gene editing using the Clustered regularly interspaced short palindromic repeats-Cas9 (CRISPR/ Cas9) system. Different delivery methods, such as microinjection (MI), electroporation (EP), and transfection (TF), can be used to transfer CRISPR/Cas9 components into porcine zygotes. This study aimed to develop a method that combines MI, EP, and TF to improve mutation efficiency mediated through the CRISPR/Cas9 system for a triple-gene knockout in pigs.

Materials and Methods: The study consisted of three groups: The MI group with three simultaneously microinjected guide RNAs (gRNAs) targeting α-1,3-galactosyltransferase (GGTA1), cytidine 32 monophosphate-N-acetylneuraminic acid hydroxylase (CMAH), and β-1,4-N-acetyl-galactosaminyltransferase 2 (B4GALNT2); the MI + EP group with two gRNAs targeting GGTA1 and B4GALNT2 genes delivered into zygotes through MI, followed by EP of gRNA targeting the CMAH 1 h later; and the MI + EP + TF group with MI of gRNA targeting GGTA1 gene into zygotes, followed by EP of gRNA targeting CMAH 1 h later, and then TF of gRNA targeting the B4GALNT2 gene into zona-free zygotes after another hour.

Results: The rate of blastocysts carrying mutations in one or two gene(s) was significantly higher in the MI + EP + TF group than in the MI group. However, the blastocyst formation rate of zygotes in the MI + EP + TF group was lower than that of the zygotes in the other treatment groups.

Conclusion: The combination of CRISPR/Cas9 delivery methods may improve the mutation efficiency of triple-gene edited porcine blastocysts.

Keywords: clustered regularly interspaced short palindromic repeats/Cas9, electroporation, microinjection, porcine zygotes, transfection.

Detection of foot-and-mouth disease virus in raw milk in Menofia Governorate and its effect on reproductive hormones and physiochemical properties of milk

Research (Published online: 16-09-2022)

11. Detection of foot-and-mouth disease virus in raw milk in Menofia Governorate and its effect on reproductive hormones and physiochemical properties of milk

Ashraf Khamees Shaban, Ragab Hassan Mohamed, Asem Mohammed Zakaria, and Eman Mohamed Baheeg

Veterinary World, 15(9): 2202-2209

ABSTRACT

Background and Aim: Foot-and-mouth disease (FMD) is an extremely contagious viral disease that affects domestic and wild cloven-hoofed animals. In Egypt, FMD has been enzootic since the 1950s and caused great economic losses in cattle and buffalos over the past few years. This study aimed to detect FMD virus (FMDV) in serum and raw milk samples collected from infected and adjacent cattle and buffalos from different localities in El Menofia Governorate, Egypt.

Materials and Methods: Blood and milk samples were collected from apparently diseased and adjacent 100 cows and 100 buffalos. Serum samples were prepared and used for the detection of FMDV using a non-structural protein enzyme-linked immunosorbent assay, while real-time reverse transcription-polymerase chain reaction (rRT-PCR) was used for the detection of FMDV in milk samples. Reproductive hormones were estimated using radioimmunoassay kits. Milk constituents were determined by Lactoscan.

Results: Of the 200 examined serum samples (100 cows and 100 buffalos), 56% and 44% were seropositive for FMDV non-structural protein antibodies in cattle and buffalo, respectively. Real-time reverse transcription-polymerase chain reaction results confirmed that all examined milk samples collected from seropositive animals were positive for FMDV. Estrogen and progesterone levels in the serum of seropositive and seronegative animals were measured, and FMDV was proven to significantly elevate estrogen and reduce progesterone levels in both non-pregnant and pregnant animals during different stages of pregnancy. The effect of the virus on milk composition and somatic cell count (SCC) was also studied, revealing that FMDV infection significantly decreased the level of milk fat, protein, and lactose but did not significantly affected minerals, pH, and conductivity. Moreover, it significantly increased the SCC.

Conclusion: Data recorded in this study indicates a widespread occurrence of FMDV in cattle and buffalo all over Menofia Governorate, Egypt. Infected raw milk is of poor quality and, if put for commercial sale, may have health risks for consumers and play a significant role in spreading the virus. Moreover, FMDV may disturb some reproductive hormones, which could adversely affect cattle and buffalo productivity. Therefore, preventive programs and accurate diagnosis are essential for successful disease control.

Keywords: foot-and-mouth disease, physiochemical properties of milk, raw milk, reproductive hormones.

Exposure to high thermal conditions for a long time induces apoptosis and decreases total RNA concentration in peripheral blood mononuclear cells among Indian Zebu–Jersey crossbreds

Research (Published online: 15-09-2022)

10. Exposure to high thermal conditions for a long time induces apoptosis and decreases total RNA concentration in peripheral blood mononuclear cells among Indian Zebu–Jersey crossbreds

Gbolabo Olaitan Onasanya, George M. Msalya, Aranganoor K. Thiruvenkadan, Nagarajan Murali, Ramasamy Saravanan, Angamuthu Raja, Moses Okpeku, Mani Jeyakumar, and Christian O. Ikeobi

Veterinary World, 15(9): 2192-2201

ABSTRACT

Background and Aim: Global warming has grave consequences on livestock production systems and profound negative effects on animal production. This study aimed to carry out an in vitro thermal stress stimulation (TSS) of bovine peripheral blood mononuclear cells (PBMCs) using different thermal assault conditions (TACs), including normal to extreme temperatures and varying durations of thermal exposure (DTE) to understand how PBMCs of Indian Zebu–Jersey crossbreds respond to various levels and durations of heat shock.

Materials and Methods: Ten milliliters of blood were collected from 70 Indian Zebu–Jersey crossbreds under aseptic conditions and were sampled for isolating PBMCs. Peripheral blood mononuclear cells were divided into seven groups, each comprising 10 PBMC samples isolated from 10 different animals. Aliquots of 500 μL of PBMCs were stressed by exposure to different TACs (37, 40, and 45°C) for DTEs of 3 or 6 h. Subsequently, the cells were harvested. The control unstressed samples (500 μL aliquots of PBMCs) were exposed to no TAC (0°C) and zero DTE (0 h). Total RNA from all the treatment groups of PBMCs were isolated and quantitated.

Results: We found a very strong association between TACs and RNA levels. In addition, PBMCs viability was negatively affected by heat shock. This led to an exponential reduction in PBMC count as TACs toughened. Only 3.59 × 105 ± 0.34 cells/mL were viable after exposure to 45°C for a 6 h DTE. This cell viability was lower than that measured in controls subjected to no stress and zero time DTE (2.56 × 107 ± 0.22 cells/mL). We also observed a reduction in the concentration of RNA isolated from thermally stressed PBMCs.

Conclusion: In vitro TSS of PBMCs provided biological information on the response of cellular systems to heat shock after exposure to TACs. This will help to mitigate and manage the effects of thermal stress in bovine species. The association between the reduction in PBMC count after in vitro TSS and the expression of heat shock protein 70 gene will be investigated in the future to further understand how Indian Zebu–Jersey crossbreds respond to in vitro thermal conditions. This will be used to determine the in vivo response of Indian Jersey crossbreds to different environmental thermal conditions and will further enable the in vivo understanding of thermotolerance potentials of bovine species for better adaptation, survival, and production performance.

Keywords: bovine, cell survival, climate change, peripheral blood mononuclear cells, RNA synthesis, thermal stress.

Genotypic characterization of mecA gene and antibiogram profile of coagulase-negative staphylococci in subclinical mastitic cows

Research (Published online: 15-09-2022)

9. Genotypic characterization of mecA gene and antibiogram profile of coagulase-negative staphylococci in subclinical mastitic cows

Eman S. Ibrahim, Sohad M. Dorgham, Asmaa S. Mansour, Abeer M. Abdalhamed, and Doaa D. Khalaf

Veterinary World, 15(9): 2186-2191

ABSTRACT

Background and Aim: Coagulase-negative staphylococci (CNS) are becoming the major cause of clinical and subclinical bovine mastitis around the world. This study aims to estimate the prevalence, antibiogram, and frequency of the methicillin-resistant (MR) (mecA) gene in CNS collected from cows with subclinical mastitis.

Materials and Methods: Thirty-four milk samples were collected from 20 cows. Fifteen subclinical mastitis samples (∼44.12%) were identified as CNS isolates. The Vitek2 compact system method was employed for the identification of the species. Furthermore, antibiotic sensitivity tests were performed against 10 different antibiotics for CNS strains. The mecA gene from isolated CNS was detected by conventional polymerase chain reaction (PCR).

Results: Staphylococcus haemolyticus was the most predominant isolated species with an incidence of 33.3% (5/15 isolates), followed by 26.7% for Staphylococcus sciuri and Staphylococcus vitamins (4/15 isolates), and 13.3% for Staphylococcus vitulinus (2/15 isolates), respectively. The highest resistance rates were determined to be 40% (6/15 isolates) against penicillin and oxacillin (OX), 33.3% (5/15 isolates) against clindamycin, 13% (2/15 isolates) against chloramphenicol, amoxicillin, and erythromycin, and 5% (1/15 isolates) against ciprofloxacin, respectively. The results revealed that the isolates were resistant to one or more antimicrobial agents, with five isolates displaying multiple antimicrobial resistance. Furthermore, the results exhibit that all CNS isolates had the mecA gene at 310 bp with a 100% frequency. Moreover, for detecting MR isolates, there are significant discrepancies between phenotypic and genotypic approaches, and only 6/15 CNS isolates phenotypically demonstrated OX resistance.

Conclusion: The results emphasize the necessity of frequent monitoring of phenotypic and genotypic profiles of CNS isolates to ensure effective control measures and the prevention of multidrug resistance strain evolution.

Keywords: coagulase-negative staphylococci, cows, mecA gene, polymerase chain reaction, subclinical mastitis.

Seroprevalence and risk assessment of Toxoplasma gondii infection in sheep and goats in North and Beqaa governorates of Lebanon

Research (Published online: 13-09-2022)

8. Seroprevalence and risk assessment of Toxoplasma gondii infection in sheep and goats in North and Beqaa governorates of Lebanon

Sara Khalife, Sara Moubayed, Rosy Mitri, Regina Geitani, and Dima El Safadi

Veterinary World, 15(9): 2180-2185

ABSTRACT

Background and Aim: Toxoplasmosis is a disease caused by the protozoan parasite Toxoplasma gondii that affects both humans and animals, leading to abortions and significant clinical manifestations in pregnant and immunocompromised hosts, in addition to massive economic losses in animal industries. Data from Lebanon are scarce regarding the seroprevalence of T. gondii infection in livestock. This study aimed to estimate the seroprevalence and assess the associated risk factors of T. gondii infection in sheep and goats in Lebanon.

Materials and Methods: A cross-sectional study was carried out from May 2020 to April 2021. Blood samples from 150 sheep and 145 goats (total 295) destined for human consumption were obtained from 20 Lebanese farms located in the North and Beqaa governorates. The anti-T. gondii immunoglobulin G antibodies were assayed through means of a modified agglutination test with a cutoff titer of 20.

Results: An overall seroprevalence of 48.5% (143/295) was reported: About 56.6% seroprevalence was found in sheep (85/150) and 40% (58/145) in goats. Adult age, female gender, and the wet season were significantly associated with an increased seropositivity rate of T. gondii infection (p < 0.001, p = 0.001, and p = 0.043, respectively).

Conclusion: These results confirm the spread of T. gondii in sheep and goats destined for human consumption in various geographical regions in Lebanon. Therefore, continuous monitoring of T. gondii infection in livestock is warranted to control the spread of the infection and limit its potential transmission to humans through the consumption of raw or undercooked meat.

Keywords: goats, Lebanon, risk factors, seroprevalence, sheep, Toxoplasma gondii.

A review: Virulence factors of Klebsiella pneumonia as emerging infection on the food chain

Review (Published online: 12-09-2022)

7. A review: Virulence factors of Klebsiella pneumonia as emerging infection on the food chain

Katty Hendriana Priscilia Riwu, Mustofa Helmi Effendi, Fedik Abdul Rantam, Aswin Rafif Khairullah, and Agus Widodo

Veterinary World, 15(9): 2172-2179

ABSTRACT

Health problems can be caused by consuming foods that have been processed in unsanitary conditions; hence, the study of the impact of contamination on food and its prevention has become critical. The disease caused by Klebsiella pneumoniae in food is increasing significantly every year across the world. The main factors that are essential for the virulence of K. pneumoniae are lipopolysaccharide and polysaccharide capsules. Furthermore, K. pneumoniae is capable of forming biofilms. Capsule polysaccharides, fimbriae types 1 and 3, are crucial virulence factors contributing to biofilm formation in K. pneumoniae. The food contamination by K. pneumoniae may not directly pose a public health risk; however, the presence of K. pneumoniae refers to unhygienic practices in food handling. This article aims to demonstrate that K. pneumoniae should be considered as a potential pathogen that spreads through the food chain and that necessary precautions should be taken in the future.

Keywords: biofilm, food chain, Klebsiella pneumonia, public health, virulence.