The Effects of Different Antimicrobial Peptides (A-11 and AP19) on Isolated Bacteria from Fresh Boar Semen and Semen Quality during Storage at 18 °C.

Antibiotic resistance (AMR) is a major public health concern. Antimicrobial peptides (AMPs) could be an alternative to conventional antibiotics. The purpose of this research was to investigate the antimicrobial ability of the synthetic AMPs (i.e., A-11 and AP19) on the most frequently isolated bacteria in boar semen and their effect on extended boar semen quality during storage. We tested the antimicrobial effect of A-11 and AP19 at different concentrations and compared them with gentamicin for inhibiting the growth of E. coli, Pseudomonas aeruginosa and Proteus mirabilis that were isolated from fresh boar semen. In order to evaluate the effect of AMP on semen qualities on days 0, 1, 3, and 5 after storage at 18 °C, seven fresh boar semen samples were collected, diluted with semen extender with antibiotic (i.e., gentamicin at 200 µg/mL, positive control) or without (negative control), and semen extender contained only A-11 or AP19 at different concentrations (i.e., 62.50, 31.25, and 15.625 µg/mL). The total bacterial count was also measured at 0, 24, 36, 48, and 72 h after storage. Comparable to gentamicin, both A-11 and AP19 inhibited the growth of E. coli, Pseudomonas aeruginosa, and Proteus mirabilis at 62.50, 31.25, and 15.625 µg/mL, respectively. Comparing the total bacterial count at 0, 24, 36, 48 and 72 h after storage, the lowest total bacterial concentration was found in the positive control group (p < 0.05), and an inferior total bacterial concentration was found in the treatment groups than in the negative control. On day 1, there is a lower percentage of all sperm parameters in the AP19 group at a concentration of 62.50 µg/mL compared with the other groups. On day 3, the highest percentage of all sperm parameters was found in the positive control and A-11 at a concentration of 31.25 µg/mL compared with the other groups. The AP19 group at 62.5 µg/mL constantly yielded inferior sperm parameters. On day 5, only A-11 at a concentration of 15.625 µg/mL showed a total motility higher than 70%, which is comparable to the positive control. A-11 and AP19 showed antimicrobial activity against E. coli, Pseudomonas aeruginosa and Proteus mirabilis isolated from boar semen. Considering their effect on semen quality during storage, these antimicrobial peptides are an alternative to conventional antibiotics used in boar semen extenders. Nevertheless, the utilization of these particular antimicrobial peptides relied on the concentration and duration of storage.

The Effect of Antimicrobial Peptide (PA-13) on Escherichia coli Carrying Antibiotic-Resistant Genes Isolated from Boar Semen.

A major global public health concern is antimicrobial resistance (AMR). Antimicrobial peptides (AMPs) are a potentially appropriate replacement for conventional antibiotics. The purpose of this research was to investigate the potential of the antimicrobial peptide PA-13, a synthetic AMP with 13 amino acids, to inhibit E. coli isolated from boar semen expressing antibiotic-resistant genes, as well as to determine the mechanism of action of this antimicrobial peptide on the bacterial membrane. The effectiveness of the bacterial inhibitory activity of PA-13 was tested at different concentrations by two fold serial dilutions in the range 0.488-500 µg/mL using the MIC and MBC methods. The impact of PA-13 on the bacterial membrane was examined at different concentrations of 0×, 0.5×, 1×, 2× and 4× of MIC using DNA leakage assay and electron microscopy. The PA-13 antibacterial activity result exhibited the same MIC and MBC values at a concentration of 15.625 µg/mL. When comparing DNA leakage at different MIC values, the results revealed that the maximum amount of DNA concentration was found two and three hours after incubation. For the results of SEM and TEM, the bacterial membrane disruption of this E. coli was found in the PA-13-treated group when compared with the negative control. In conclusion, synthetic PA-13 with its antibacterial properties is an alternative antimicrobial peptide to antibiotics in the pig industry.

Antimicrobial activity of cell free supernatants from probiotics inhibits against pathogenic bacteria isolated from fresh boar semen.

The use of antibiotics with semen extender appears to be a practical solution to minimise bacterial growth in fresh boar semen preservation. Unfortunately, the excessive use of antibiotics promotes antimicrobial resistance (AMR). This becomes a worldwide concern due to the antimicrobial resistance genes transmitted to animals, environment, and humans. Probiotics are one of the alternative methods to reduce antibiotic use. They could inhibit pathogenic bacteria by producing antimicrobial substances in cell free supernatants (CFS). Nevertheless, there is no comprehensive study undertaken on inhibitory activity against pathogenic bacteria isolated from boar semen origin. Our study investigated the efficacy of CFS produced from selected probiotics: Bacillus spp., Enterococcus spp., Weissella spp., Lactobacillus spp., and Pediococcus spp. inhibiting pathogenic bacteria isolated from fresh boar semen. Besides, the semen-origin pathogenic bacteria are subjected to identification, antimicrobial resistance genes detection, and antibiotic susceptibility test (AST). Pseudomonas aeruginosa, Escherichia coli, and Proteus mirabilis are the most common pathogens identified in boar semen with resistance to numerous antibiotics used in pig industry. The CFS with its antimicrobial peptides and/or bacteriocin constituent derived from selected probiotics could inhibit the growth of pathogenic bacteria carrying antimicrobial resistance genes (mcr-3 and int1 genes). The inhibition zones for Pseudomonas aeruginosa, Escherichia coli, and Proteus mirabilis provided more efficient results in the CFS derived from Lactobacillus spp. and Pediococcus spp. than those of the CFS produced from Enterococcus spp., Weissella spp. and Bacillus spp., respectively. It is worth noted that as the incubation time increased, the antibacterial activity decreased conversely. Our results on CFS with its antimicrobial peptides and/or bacteriocin constituent inhibits semen-origin pathogenic bacteria guide the direction as a promising alternative method used in the semen extender preservation of the pig industry.

Antibiotic resistant Escherichia coli from diarrheic piglets from pig farms in Thailand that harbor colistin-resistant mcr genes.

Antibiotic-resistant Escherichia coli is one of the most serious problems in pig production. This study aimed to determine the antibiotic susceptibility and genotypes profiles of diarrhoeagenic E. coli that causes diarrhea in piglets. Thirty-seven pathogenic E. coli strains were used in this study. These were isolated from rectal swabs of diarrheic piglets from farms in Thailand from 2018 to 2019. Escherichia coli isolates were highly resistant to amoxicillin (100%), followed by oxytetracycline (91.9%), enrofloxacin (89.2%), trimethoprim/sulfamethoxazole (86.5%), amoxicillin: clavulanic acid (81.1%), colistin and gentamicin (75.7%), ceftriaxone and ceftiofur (64.9%), ceftazidime (35.1%) and 97.3% showed multidrug-resistance (MDR). There were 8 (21.6%) mcr-1 carriers, 10 (27.0%) mcr-3 carriers and 10 (27.0%) co-occurrent mcr-1 and mcr-3 isolates. The phenotype-genotype correlation of colistin resistance was statistically significant (performed using Cohen's kappa coefficient (κ = 0.853; p < 0.001)). In addition, PCR results determined that 28 of 37 (75.7%) isolates carried the int1 gene, and 85.7% int1-positive isolates also carried the mcr gene. Genetic profiling of E. coli isolates performed by ERIC-PCR showed diverse genetics, differentiated into thirteen groups with 65% similarity. Knowledge of the molecular origins of multidrug-resistant E. coli should be helpful for when attempting to utilize antibiotics in the pig industry. In terms of public health awareness, the possibility of transmitting antibiotic-resistant E. coli from diarrheic piglets to other bacteria in pigs and humans should be of concern.