BIO Web Conf.
Volume 33, 2021The 1st International Conference of Advanced Veterinary Science and Technologies for Sustainable Development (ICAVESS 2021)
|Number of page(s)||5|
|Section||Zoonosis, Emerging Disease, and Transboundary Animal Disease|
|Published online||23 August 2021|
Molecular Screening of Salmonella sp. from fecal sample of Sparrows (Passer domesticus) in Yogyakarta, Indonesia
Department of Microbiology, Faculty of Veterinary Medicine, Universitas Gadjah Mada, Yogyakarta 55281, Indonesia
* Corresponding author: Marla.firstname.lastname@example.org
Wild birds is one of the reservoir agent of some of various zoonotic diseases. The study was aim to see the potential of sparrow as the reservoir agent of Salmonella sp. using polymerase chain reaction (PCR) method. We detected the invA gene of Salmonella sp. from faecal sample of sparrows (Passer domesticus) in local area of Yogyakarta, Indonesia. A total of 30 faecal dropping samples were collected from sparrows. DNA was extracted from the faecal samples, then amplified by PCR for the target genes. The amplicons were electrophorized to see the visualization of DNA on the agarose gel. The result showed the prevalence of the positive result of Salmonella sp. was 3,3%. The study indicated that sparrows can spread zoonotic pathogens and this necessitates monitoring for the epidemiologic status of these pathogens among birds, also applying the appropriate intervention measures to prevent the transmission of zoonotic diseasesfrom birds to humans.
© The Authors, published by EDP Sciences, 2021
This is an Open Access article distributed under the terms of the Creative Commons Attribution License 4.0, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
The current pandemic issues in the world currently has made public attention to the various possibilities of wild animals as the important objects in the spread of zoonotic diseases. In addition to mamals, wild birds are also considered as an important reservoir of some pathogens zoonoses . Several species of wild birds are known to act as reservoir for zoonotic agents such as Chlamydophila psittaci , Avian Influenza , α-, β-, γ-, and δcoronaviruses  enteropathogenic E.coli (EPEC) and Shiga-toxin producing E. coli (STEC) , and Salmonella . The transmission of the diseases occurs between the wild birds to other animals and humans through droplets of feces or nasal fluid [7, 8]. Salmonella sp. spreaded through direct contact with bacteria-contaminated feces [9, 10]. Type of wild birds included in the order Passeriformes, Psittaciformes, and Columbiformes are recorded as a reservoir of various zoonotic diseases caused by bacteria [11, 12].
Salmonella enterica is one of the important member of enterobacteriaceae family. Salmonella enterica serovar Typhimurium (S. Typhimurium) is responsible for acute gastroenteritis or typhoid fever in humans , acute enteritis in cattle and pigs, S. enterica serovar Gallinarum (S. Gallinarum) and Pullorum (S. Pullorum), the cause of fowl typhoid and pullorum disease, respectively diseases in avian . Diagnosis of Salmonella can be carried out by culture of feces, blood, spleen, liver, and intestinal contents. In addition, very small numbers of viable organisms present in the feces may fail to grow in artificial laboratory media [15, 16]. Recently, the molecular techniques like polymerase chain reaction (PCR), DNA microarray-based detection, and DNA sequencing were developed and used to detect bacterial infection in different clinical materials [17, 18]. Molecular testing has been most successful in areas for which conventional microbiologic techniques do not exist, are too slow, or are too expensive .
This study was aim to detect Salmonella sp. from fecal sample of the sparrow birds, and to know the prevalence of Sparrows as vectors of Salmonella sp. in Yogyakarta area, Indonesia.
This study has the ethical requirements for research in experimental animals from the Integrated Research and Testing Laboratory (LPPT) of Gadjah Mada University with certificate number: 00033/04/LPPT/VI/2019. Thirty sparrow birds were obtained from the bird hunter in Yogyakarta. Fresh fecal samples were collected and Genomic DNA was extracted using Stool DNA Isolation Mini Kit (Favorgen, Biotech Corp).
Salmonella sp. detection was carried out using invA gene targeted primers forward (5’CGGTGGTTTTAAGCGTACTCTT-3’) and reverse (5’-CGAATATGCTCCACAAGGTT A-3’) with the invA gene target (769 bp).
The PCR reaction for invA gene of Salmonella sp. was carried out under initial denaturation conditions at a temperature of 94 °C for 2 mins, followed by 38 cycles consisting of denaturation at a temperature of 94 °C for 20 s , annealing at 60 °C for 1 min, and extension at 72 °C for 1 minute. The PCR products were electrophorized to see the amplified DNA band. Electrophoresis was performed at 100V for 30 minutes. The gel then visualized under UV light to see the DNA band.
This study reports the prevalence of Salmonella sp. in sparrow birds from wild of Yogyakarta region, Indonesia from the fecal samples. Fecal swab samples performed PCR using invA genes showed one positive sample of Salmonella sp (3.3%) (Figure 1). Salmonella sp. detection was carried out using invA gene.
According to Malorny et al.  the invA gene became an international standard for detecting Salmonella sp. at the genus level using clinical samples such as feces. The percentage of Salmonella sp. in sparrows in Yogyakarta is higher when compared to previous studies in various countries except UK, which has a percentage of Salmonella sp in wild birds of 1.6% . Research conducted by Afema & Sisco  found that the percentage of Salmonella sp in birds was 4.1%, this is higher than the percentage shown in this study. The same result was revealed by Tizard , who found the prevalence of Salmonella sp in captive raptors was 7.36%, higher than the prevalence of Salmonella in sparrows in Yogyakarta. Sparrows as reservoir for Salmonella sp was expressed as carriage with no obvious disease manifestations. Some family of passerines are the carriers of Salmonella strains. In the study by Krawiec et al. , they discovered Salmonella serovar Typhimurium positive samples from Eurasian siskins and greenfinches. Salmonella can infect wild birds through direct contact with the vectors such as insects and rodents, or through food-producing animals .
Free-living birds as well as migratory and captivated birds, are potential as reservoirs for bacterial agents . They also may act as vectors in the transmission of some pathogens . Matias et al.  stated that there are relevance in illegal trading of wild birds with outbreaks potential of some zoonotic diseases in human. Environmental changes, forest loss, human demographic changes, industry expand, could lead direct contact between wild animals and humans. There is also the possibilities of the increase risk of pathogen spread from the wild animals to farm animals, thus resulting in foodborne diseases to humans. The interface between livestock and wildlife has been reported associated with rising incidences of pathogens . Safety measure for humans at high risk of zoonotic infection from livestock and wild animals must be taken to prevent the occurrence of outbreak in humans.
Polymerase chain reaction product amplifying invA gene of Salmonella sp. (arrow) on agarose gel electrophoresis. M: marker, K (-): negative control, 2-15: samples
This study showed the prevalence of Salmonella sp. from the fecal sample of wild sparrows in Yogyakarta was 3.3%. The sparrows can act as a carrier by spreading the bacteria through feces without showing any clinical symptoms. Further research on the pathogens in wild birds, that are abundant in Indonesia, especially in Yogyakarta region is needed to determine variations in the types of zoonotic diseases that can be carried by wild birds. Evaluation and monitoring of the epidemiologic status of these pathogens among birds and humans is needed for applying the appropriate intervention measures to prevent the transmission of zoonotic diseases.
This study was supported by The Research Fund of Faculty of Veterinary Medicine, Universitas Gadjah Mada (BPPTN-BH FKH UGM), Indonesia with grant number: /J01.1.22/HK4/2019.
- M. Wille and C. Holmes. Wild birds as reservoirs for diverse and abundantgamma-and deltacoronaviruses. FEMS Microbiology Reviews, fuaa026, 44:631-644 (2020) [Google Scholar]
- T. Piasecki, K. Chrzastek and A. Wieliczko. Detection and identification of Chlamydophila psittaci in asymptomatic parrots in Poland. Piasecki et al. BMC Veterinary Research, 8:233 (2012) [Google Scholar]
- M. Richard, R. Fouchier, I. Monne, and T. Kuiken. Mechanism and risk factors for mutation from low to highly pathogenic avian influenza virus. EFSA supporting publication 2017: EN-1287. 26 pp. doi:10.2903/sp.efsa.2017.EN-1287 (2017) [Google Scholar]
- H. Hu, K. Jung, Q. Wang, L.J. Saif, and A.N. Vlasova. Development of a one-step RT PCR assay for detection of pancoronaviruses (α-, β-, γ-, and δ-coronaviruses) using newly designed degenerate primers for porcine and avian fecal samples, Journal of Virological Methods https://doi.org/10.1016/j.jviromet.2018.02.021 (2010) [Google Scholar]
- L.A. Sanches, M. Gomes da Silva, R.H.F. Teixeira, M.P.V. Cunha, M.G.X. Oliveira, M.A.M. Vieria, T.A.T. Gomes, and T. Knobl. Captive wild birds as reservoirs of enteropathogenic E. coli (EPEC) and Shiga-toxin producing E. coli (STEC). Brazilian journal of microbiology, 48:760-763 (2017) [Google Scholar]
- M.P. Stevens, T.J. Humphrey, and D.J. Maskell. Molecular insights into farmanimal and zoonotic Salmonella infectons. Phil. Trans. R. Soc. B., 364:2709-2723 (2009) [Google Scholar]
- J. F. W. Chan, K. K. W. To, H. Chen, and K. Y. Yuen. Cross-species transmission and emergence of novel viruses from birds. Current opinion in virology, 10:63-69 [Google Scholar]
- H. Gerlach. Chlamydia. In: Avian medicine: Principles and Application. HBD International Inc., Delray Beach, Florida. 984-996 (1999) [Google Scholar]
- B. Sareyyupoglu, A. Celik Ok, Z. Cantekin, H. Yardimci, M. Akan, and A. Akcay. Polymerase Chain Reaction Detection of Salmonella spp. in Fecal Samples ofPetBirds. Avian Diseases, 52(1): 163-167 (2007) [Google Scholar]
- T. Harkinezhad, T. Geens, and D. Vanrompany. Chlamydophila psittaci Infection in Birds: A Review with emphasis on zonotic consequences. Vet Microbiology 135 (1): 68-77 (2009) [Google Scholar]
- G. Boseret, B. Losson, J.G. Mainil, G. Thiry, C. Saegerman. Zoonoses in pet birds: review and perspectives. Veterinary Research 2013, 44:36 (2013) [Google Scholar]
- M. Krawiec, T. Piasecki, and A. Wieliczko. Prevalance of Chlamydia psittaci and other Chlamydia Species in Wild Birds in Poland. Vector Borne and Zoonotic Diseases Vol 15 No 11: 652-655 (2015) [Google Scholar]
- C.V. Srikanth and B.J. Cherayll. Intestinal Innate Immunity and the Pathogenesis of Salmonella Enteritis. Immunol Res, 37(1):61-78 (2007) [Google Scholar]
- D. M. Heithoff, W. R. Shimp, P. W. Lau, G. Badie, E. Y. Enioutina, R. A. Daynes, B. A. Byrne, J. K. House, and M. J. Mahan. Human Salmonella clinical isolates distinct from those of animal origin. Appl. Environ. Microbiol. 74, 1757–1766. (doi:10.1128/AEM.m02740-07) (2008) [Google Scholar]
- I.S. Schrank, M.A.Z. Mores, J.L.A. Costa, A.P.G. Frazzon, R. Soncini, A. Schrank, M.H. Vainstein, and S.C. Silva. Influence of enrichment media and application ofaPCR based method to detect Salmonella in poultry industry products and clinical samples. Vet. Microbiol. 82:45-53 (2011) [Google Scholar]
- I. Feder, J.C. Nietfeld, J. Galland, T. Yeary, J.M. Sargeant, R. Oberst, M.L. Tamplin, and J.B. Luchansky. Comparison of cultivation and PCR-hybridization for detectionof Salmonella in porcine fecal and water samples. J. Clin. Microbiol. 39:2477-2484 (2001) [Google Scholar]
- S.D. Oliveira, L.R. Santos, D.M.T. Schuch, A.B. Silva, C.T.P. Salle, and C.W. Canal. Detection and identification of salmonellas frompoultry-related samples by PCR. Vet Microbiol. 82:45-53 (2002) [Google Scholar]
- P. Whyte, K. McGill, J.D. Collins, and E. Gormley. The prevalence and PCR detection of Salmonella contamination in raw poultry. Vet. Microbiol. 89:53-60 (2002) [Google Scholar]
- V.K. Sharma, and S.A. Carlson. Simultaneous detection of Salmonella strains and Eschericia coli O157:H7 with fluorogenic PCR and single-enrichment-broth culture. Appl. Environ. Microbiol. 66:5472-5476 (2000) [Google Scholar]
- B. Malorny, J. Hoorfar, M. Hugas, A. Heuvelink, P. Fach, L. Ellerbroek, C. Bunge, C. Dorn, and R. Helmut. Inter laboratory diagnostic accuracy of a Salmonella specific PCRbased method. Int. J. Food Microbiol., 89: 241-249 (2003) [Google Scholar]
- B. Lawson, E. Pinna, R.A. Horton, S.K. Macgregor, S.K. John, J. Chantrey, P.J. Duff, J.K. Kirkwood, V.R. Simpson, R.A. Robinson, J. Wain, A.A. Cunningham. Epidemiological Evidence That Garden Birds are a Source of Human Salmonellosis in England and Wales. Plos One 9 (2) (2014) [Google Scholar]
- J.A. Afema and W.M. Sischo. Salmonella in Wild Birds Utilizing Protected andHuman Impacted Habitats, Uganda. EcoHealth 13:558-569 (2016) [Google Scholar]
- I. Tizard. Salmonellosis in wild birds. Seminars in Avian and Exotic Pet Medicine, 13(2):50–66 (2004) [Google Scholar]
- M. Krawiec, M. Kuczkowski, A.G. Kruszewicz, and A. Wieliczko. Prevalence and genetic characteristics of Salmonella in free-living birds in Poland. BMC Veterinary Research 11:15 (2015) [Google Scholar]
- F. Hilbert, F. J. M. Smulders, R. Chopra-Dewasthaly, and P. Paulsen, “Salmonellainthe wildlife-human interface, ” Food Research International, 45(2): 603–608 (2012) [Google Scholar]
- M. Foti, D. Rinaldo, A. Guerci, C. Giacopello, A. Aleo, F. De Leo, et al. Pathogenic microorganisms carried by migratory birds passing through the territory oftheislandof Ustica, Sicily (Italy). Avian Pathol. 40:405–9. (2011) [Google Scholar]
- C. M. H. Benskin, K. Wilson, K. Jones, and I. R. Hartley, “Bacterial pathogens in wild birds: a review of the frequency and effects of infection, ” Biological Reviews, 84(3):349–373 (2009) [Google Scholar]
- C. A. R. M. Matias, Pereira I. A., Araujo M. Dos S., Santos A. F. M., Lopes R. P., Christakis S., Rodrigues D. Dos P., and Siciliano S. Characteristics of Salmonella spp. Isolated from Wild Birds Confiscated in Illegal Trade Markets, Rio de Janeiro, Brazil. BioMed Research International, 2016, Article ID 3416864, 7 pages. http://dx.doi.org/10.1155/2016/3416864 (2016) [Google Scholar]
- B. A. Jones, D. Grace, R. Kock, S Alonso., J. Rushton, M. Y. Said, et al. Zoonosis emergence linked to agricultural intensification and environmental change. Proc Natl Acad Sci USA. 110:8399 404. doi: 10.1073/pnas.1208059110. (2013) [Google Scholar]
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