Antimicrobial susceptibility test and antimicrobial resistance gene detection of extracellular enzyme bacteria isolated from tilapia (Oreochromis niloticus) for probiotic candidates

Research (Published online: 11-02-2023)

4. Antimicrobial susceptibility test and antimicrobial resistance gene detection of extracellular enzyme bacteria isolated from tilapia (Oreochromis niloticus) for probiotic candidates

Mira Mawardi, Agustin Indrawati, I. Wayan Teguh Wibawan, and Angela Mariana Lusiastuti

Veterinary World, 16(2): 264-271

ABSTRACT

Background and Aim: Antimicrobial resistance (AMR) is a global problem that can increase mortality and morbidity rates and adversely affect health. Therefore, AMR control must be carried out in various sectors, including the fisheries sector, using probiotics. Bacteria can become resistant to antibiotics, including bacteria used for probiotics. This study aimed to isolate bacteria as potential producers of extracellular enzymes, phenotypic characterization, and antibiotic-resistant gene patterns.

Materials and Methods: In this study, 459 bacterial isolates were isolated from the stomach of tilapia in Indonesia. Tilapia was obtained from Sukabumi, Ciamis, Serang, Banjarnegara, Jayapura, Sorong, Manokwari Selatan, Takalar, Lampung, Batam, and Mandiangin. Enzymatic bacteria were identified. An antimicrobial susceptibility test was conducted by agar disk diffusion, and genotypic detection of encoding genes was performed using a molecular method.

Results: This study obtained 137 isolates (29.84%) that can produce extracellular enzymes. The highest number of E-sensitive isolates was found, including 130 isolates (94.89%). Six isolates (6/137) can produce four enzymes (amylase, protease, cellulose, and lipase), and they were sensitive to antibiotics. A total of 99 isolates can produce extracellular enzymes, and they were sensitive to antibiotics. Such isolates serve as a consortium of probiotic candidates. The isolates that are resistant to oxytetracycline (OT), erythromycin (E), tetracycline (TE), and enrofloxacin (ENR) included 15 isolates (10.95%), seven isolates (5.11%), three isolates (2.19%), and one isolate (0.73%), respectively. In addition, four isolates (2.92%) were detected as multidrug-resistant. The tet(A) gene obtained the highest result of detection of resistance genes in isolates that were intermediate and resistant to TE and OT. Isolates that serve as ENR intermediates have a high qnr(S) resistance gene.

Conclusion: The data in this study provide the latest update that bacteria can serve as a consortium of potential probiotics with antibiotic-resistant genes for the treatment of fish. Bacteria that are intermediate to antibiotics may contain resistance genes. The results of this study will improve the policy of probiotic standards in Indonesia.

Keywords: antibiotic, antimicrobial resistance, enzymatic bacteria, Oreochromis niloticus, probiotic.

Effect of claw blocks on the healing duration and lesion severities of claw lesions in lame cows in Western Thailand

Research (Published online: 11-02-2023)

3. Effect of claw blocks on the healing duration and lesion severities of claw lesions in lame cows in Western Thailand

Pipat Arunvipas, Teerachad Setkit, Jaturong Wongsanit, Theera Rukkwamsuk, Nitipong Homwong, and Anawat Sangmalee

Veterinary World, 16(2): 258-263

ABSTRACT

Background and Aim: Lameness is a major complication in dairy cattle affecting health and milk production. Several factors are found to contribute to this condition and specific treatments are required, including the process of claw trimming. The elevation of the claw, such as with the application of a claw block, was reported to be beneficial in the more severe cases. This study aimed to determine the efficiency of a claw block on claw lesions of lame cows in dairy farms in Western Thailand.

Materials and Methods: Locomotion scores of 376 dairy cows were determined by a veterinarian using a scale of 1–5 (1 = normal; 5 = severely lame) at the time of the visit. Cows with a score of 3 or greater were defined as clinically lame. In total, 134 clinically lame cows from 11 dairy farms in the Kanchanaburi and Ratchaburi provinces were included in the analysis. Claw lesions included a white line abscess, bruised sole, sole ulcer, sole abscess, white line separate, and double soles. Wooden or rubber claw blocks were applied to the unaffected claw of the same hoof as the injured claw of 116 cows, which were classified as the treatment cases, and 18 cows were left untreated and classified as the control cases. Each cow was checked on every week of the healing process for 2 months unless the cow was culled earlier. Survival analysis was based on the Kaplan–Meier estimator and Cox Proportional Hazard regression.

Results: The median healing time for lame cows with and without claw blocks was 21 and 36 days, respectively. After adjusting for the lesion severity and type, the lame cows with and without a claw block had hazard ratios of 2.16 and 3.08, respectively. The healing times between the four lesion types in cows with a claw block were not significantly different. The healing time was longer in lame cows, with a severity score of 4.

Conclusion: The results from this study reveal that the treatment of lame cows with claw blocks promoted the healing capacity of claw lesions after claw trimming.

Keywords: claw block, claw lesion, dairy cows, lameness.

Aeromonas hydrophila induction method in adult zebrafish (Danio rerio) as animal infection models

Research (Published online: 09-02-2023)

2. Aeromonas hydrophila induction method in adult zebrafish (Danio rerio) as animal infection models

Dahliatul Qosimah, Sanarto Santoso, Maftuch Maftuch, Husnul Khotimah, Loeki Enggar Fitri, Aulanni'am Aulanni'am, and Lucia Tri Suwanti

Veterinary World, 16(2): 250-257

ABSTRACT

Background and Aim: Zebrafish are frequently used as model organisms in scientific research as their genes mirror those of humans. Aeromonas hydrophila bacteria can infect humans and animals, mainly fish. This study aimed to identify the concentration and route of A. hydrophila infection in adult zebrafish. Zebrafish had been used as a challenge test by analyzing their hematological profiles, blood glucose levels, and survival rates.

Materials and Methods: Induction of cell supernatant free (CSF) from A. hydrophila bacteria in adult zebrafish was carried out via bath immersion (BI), intraperitoneal injection (IPI), intramuscular injection (IMI), and healthy zebrafish as a control (C). The bacterial concentrations were 107, 109, and 1011 colony-forming units (CFU)/mL. At 24 h post-infection, the outcomes of infection were evaluated based on survival rates, hematological profiles, and blood glucose levels. A one-way analysis of variance with a confidence level of 95% was employed to examine the data.

Results: In the BI, IPI, and IMI treatment groups, the survival rate of the fish reached a peak of 100%, 22%–100%, and 16%–63%, respectively, compared with the injection technique. In the IMI2 group, a 109 CFU/mL bacterial concentration was determined to correspond to the lethal dosage 50. All infection groups had lower erythrocyte and hemoglobin counts but higher leukocyte counts than the control group. The blood sugar levels of the healthy and infected groups were not significantly different.

Conclusion: The route of A. hydrophila infection through Intramuscular injection with a concentration of 109 CFU/mL indicated a high performance compared to other techniques. This method could be developed as a reproducible challenge test.

Keywords: Aeromonas hydrophila, animal model, hematological profile, survival rate, zebrafish.

Veterinary World reviewer acknowledgment 2022

Reviewer Acknowledgment 2022 (Published online: 08-02-2023)

1. Veterinary World reviewer acknowledgment 2022

A. V. Sherasiya and Nazir

Veterinary World, 16(2): 246-249

Contributing reviewers 

Veterinary World editorial team would sincerely like to thank all of our reviewers who contributed to peer review for the journal in 2022.

Methicillin-resistant Staphylococcus aureus isolates derived from humans and animals in Yogyakarta, Indonesia

Research (Published online: 31-01-2023)

29. Methicillin-resistant Staphylococcus aureus isolates derived from humans and animals in Yogyakarta, Indonesia

Mulya Fitranda, Siti Isrina Oktavia Salasia, Osman Sianipar, Dion Adiriesta Dewananda, Adika Zhulhi Arjana, Fatkhanuddin Aziz, Madarina Wasissa, Fajar Budi Lestari, and Christin Marganingsih Santosa

Veterinary World, 16(1): 239-245

ABSTRACT

Background and Aim: The emergence of methicillin-resistant Staphylococcus aureus (MRSA) as a highly pathogenic strain in veterinary and human medicine is a growing global problem. This study aimed to evaluate MRSA isolates of human and animal origin against various antibiotics in Yogyakarta, Indonesia.

Materials and Methods: The susceptibility test was carried out by the disk diffusion method using Mueller-Hinton agar against nine antibiotic disks. Methicillin-resistant S. aureus strains were genetically confirmed through mecA gene detection encoding for methicillin resistance by polymerase chain reaction.

Results: All 240 S. aureus strains isolated from animals and humans were resistant to penicillin G (P) (100% and 99%, respectively), followed by ampicillin (AMP), amoxicillin (AML), oxacillin (OX), erythromycin (E), clindamycin (DA), tetracycline (TE), gentamicin (GEN), and ciprofloxacin (CIP). Eighty-three MRSA strains were resistant to OX (100%), P (100%), AMP (99.27%), AML (95.52%), E (87.77%), TE (71.33%), DA (63.24%), GEN (38.81%), and CIP (26.87%).

Conclusion: The antimicrobial resistance pattern of S. aureus human isolates was similar to their animal counterpart, with 77.20% of MRSA strains classified as multidrug-resistant (MDR) bacteria. These findings indicate an increase in MDR S. aureus strains of animal origin in Yogyakarta, thus raising public health concerns about MRSA zoonotic spread.

Keywords: antimicrobial resistance, methicillin-resistant Staphylococcus aureus, multi-drug resistance, Staphylococcus aureus.

First seroprevalence and molecular identification report of Brucella canis among dogs in Greater Cairo region and Damietta Governorate of Egypt

Research (Published online: 31-01-2023)

28. First seroprevalence and molecular identification report of Brucella canis among dogs in Greater Cairo region and Damietta Governorate of Egypt

Mahmoud E. R. Hamdy, Mahmoud H. Abdel-Haleem, Rehab E. Dawod, Rania I. Ismail, Soliman S. Hazem, Hanan A. Fahmy, and Nour H. Abdel-Hamid

Veterinary World, 16(1): 229-238

ABSTRACT

Background and Aim: Given the rise in stray and imported dogs in Egypt over the past 5 years, it is surprising that no report of Brucella canis infection in dogs or humans has been documented in Egypt's published papers. This study aimed to detect the presence of antibodies against the rough (B. canis) and smooth Brucellae among dogs in Egypt and to characterize the Brucella species circulating in dogs.

Materials and Methods: Blood samples (n = 449) were collected from owned and stray dogs in the Greater Cairo region (n = 309) and Damietta governorate (n = 140). The apparent, true, and total seroprevalence of canine brucellosis caused by B. canis infection were calculated using the 2-mercaptoethanol tube agglutination test (2-ME TAT) and rapid slide agglutination test (RSAT). We used the rose Bengal test (RBT) and the buffered acidified plate antigen test (BAPAT) to check the serum samples from dogs for the presence of antibodies against smooth Brucellae. Three polymerase chain reaction (PCR) assays - Bruce-ladder PCR, B. canis species-specific PCR (BcSS-PCR), and Abortus Melitensis Ovis Suis (AMOS)-PCR - were used to determine the Brucella species in the buffy coats of the serologically positive dogs.

Results: The overall apparent and true prevalence of B. canis infection in dogs were estimated to be 3.8% and 13.2%. The estimated true prevalence in stray dogs (15%) was higher than in owned dogs (12.5%). The BAPAT and the RBT using smooth antigens revealed that 11 (2.4%) and 9 (2%) were positive. Bruce-ladder PCR targeting eryC, ABC, and Polysaccharide deacetylase genes was able to identify B. canis in nine out of 17 buffy coat samples. AMOS-PCR identified the eight undetermined Brucella species by Bruce-ladder PCR as Brucella abortus (n = 4) and Brucella melitensis (n = 4). To exclude the presence of Brucella suis, a one-step species-specific BcSS-PCR was performed and specifically amplified all B. canis DNA (n = 9) the same as did the Bruce-ladder PCR.

Conclusion: To the best of our knowledge, this is the first report of B. canis detection in dogs in Egypt. Molecular identification of B. abortus and B. melitensis in the Egyptian canines highlights the role of stray dogs in brucellosis remerging in Brucellosis-free dairy farms. Brucella canis infection can be diagnosed specifically with the one-step BcSS-PCR. The obtained results set-an-alarm to the veterinary authorities to launch plans to control this disease in dogs.

Keywords: 2-mercaptoethanol tube agglutination test, AMOS-polymerase chain reaction, Bruce-ladder polymerase chain reaction, Brucella canis species-specific-polymerase chain reaction, Brucella canis, rapid slide agglutination test.

Expression of recombinant Omp18 and MOMP of Campylobacter jejuni and the determination of their suitability as antigens for serological diagnosis of campylobacteriosis in animals

Research (Published online: 30-01-2023)

27. Expression of recombinant Omp18 and MOMP of Campylobacter jejuni and the determination of their suitability as antigens for serological diagnosis of campylobacteriosis in animals

Sergey Borovikov, Kanat Tursunov, Alfiya Syzdykova, Ainagul Begenova, and Alfira Zhakhina

Veterinary World, 16(1): 222-228

ABSTRACT

Background and Aim: Campylobacteriosis causes gastrointestinal tract lesions in adults and children and may result in severe complications. The primary sources of infection are infected animals and animal products. Immunochemical methods effectively diagnose intestinal infections but require highly specific antigens to detect their antibodies. This study aimed to obtain two recombinant immunogenic antigens of Campylobacter jejuni, an outer membrane protein with a molecular weight of 18 kDa (Omp18) and the major outer membrane protein (MOMP) with a molecular weight of 45 kDa, and evaluate their suitability for the serological diagnosis of campylobacteriosis using immunochromatographic assay (ICA).

Materials and Methods: The C. jejuni Omp18 and MOMP gene sequences were synthesized de novo (Macrogen, Korea) and cloned into the pET32 expression plasmid. Using these genetic constructs, electrocompetent cells of the Escherichia coli BL21 strain were transformed and cultured under various conditions. Antigens were purified and refolded using metal affinity chromatography. The properties of the purified proteins were studied by western blotting, liquid chromatography with tandem mass spectrometry, and enzyme-linked immunosorbent assay (ELISA).

Results: We developed two recombinant E. coli BL21 cells producing rOmp18 and Recombinant MOMP (rMOMP) antigens with molecular weights of 36 and 64 kDa, respectively. Amino acid sequence analysis of the obtained antigens showed complete homology with the reference sequences in the PubMed NCBI database. Western blotting using positive-control sera demonstrated the specificity of the recombinant antigens. The results of ELISA with 94 bovine sera showed the interaction of recombinant antigens with specific antibodies.

Conclusion: The obtained rOmp18 and rMOMP antigens can detect antibodies in the serum of infected or recovered animals and can be used to develop ICA.

Keywords: Campylobacter jejuni, campylobacteriosis, diagnostics, outer membrane proteins, recombinant antigens.