Profil dan gen resistensi antimikroba Salmonella Enteritidis asal ayam, telur, dan manusia

(1)

PROPIL DAN GEN RESISTENSI ANTIMIKROBA

Salmonella

Enteritidis ASAL AYAM, TELUR,

DAN MANUSIA

ANN1 KUSUMANINGSIH

SEKOLAH PASCASARJANA

INSTITUT

. .

PERTANIAN

BOGOR

. .

BOGOR

.

(2)

PERNYATAAN

MENGENAI DISERTASI

DAN

SUMBER

INFORMASI

Dengan ini saya menyatakan bahwa disertasi Profil l Gen Resistensi

Antimikroba Salmonella Enteritidis Asal Ayam, Telur, dan Mmwia adalah karya

saya sendiri dengan arahan komisi pembirnbing

dan belum pernah diajukan dalarn

berituk apapun kepada perguruan tinggi mana pun. Sumber informasi yang berasal

atau d i t i p dari karya yang ditubitkan

dm

tidak diierbitkan

dari

_penulis_lain

(3)

ABSTRACT

ANNI

KUSUMANINGSIH.

The Profile and Genes of Antimicrobes Resistance of Safmoneffa Enteritidis from Chickens, Eggs, and Human. Supervised by Hj. MIRNAWATI SUDARWANTO, I WAYAN TEGUH WIBAWAN, ROCHMAN NA'IM and SUPAR

SalmoneIlosis associated with Salmonella Enteritidis infection is considered

as

an important disease in poultry industries. The disease in chicken sometimes shows no clinical signs, but the agents persist in the lymphatic tissues and contaminated their produck The infection in human due to consume chickens, eggs, and theirs products contaminated

with

S. Enteritidis. The objectives of this study were: to produce

0

somatic (9) and

H

flagella

(m)

antisera for serotyping; to obtain

S.

Enteritidis isolates from chickens, eggs, and human origin; to study the antimicrobial resistance profile of S. Enteritidis isolates; to observed the number and size of plasmid from S. Enteritidis; and to detect the resistance genes of tetracycline, chloramphenicol, and trimetoprim- sulphametoxazol from S. Enteritidis plasmids and chromosomes. Samples of chicken cloaca1 swabs, livers and hearts, and eggs were collected h r n _{farms and}

markets

in Bogor, Sukabumi, Cianjur, and Bandung; swab samples f b m human were also collected from hospital; some Salmonella spp. isolates were obtained fiom Laboratory of Microbiology, Medical School,

The

University of Indonesia, Jakarta; and private laboratory in Bogor. Thirty three isolates of S. Enteritidis fiom Balitvet Culture Colection werc: included in this study. Antimicrobe sensitivity assays of 68 S. Enteritidis indicated that 90.1% of these isolates were isolated from chickens, 14 isolates fiom eggs, and 13 isolates h m human. Sixty three of 68 (90,10%) S. Enteritidis isolates were resistance to streptomycin, neornycin, gentamycin, chloramphenicol, tetracycline, oxytetracycline, doxicycline, trimetophrim, ciproflocacin, and enrofloxacin-The multiresistance

between

2 to 9 antimicrobial agents: such as streptomycin, neomicyn, gentamicin tetrrrcyclin, doxicycline, and ciprofloxacine. Twenty five isolates of S. Enteritidis fiom chkken, eggs, and 'hLuriar1 tested qualitatively against chloramphenicol, tetracycline,

-tdmetophrim-sulhpmetoxazol,

and ciprofloxacin demonstrated MICs - of betwe"e6 12.50-50.0 pg/rnl, 0.8-50.0 pglml, 6.3-50.0 pg/ml, and less than 0.1 pgml respectively. Twenty one of 25 (84%) isolates which were resistant to 10 different antimicrobes carried 1-6 plasmids with differences in sizes: 0.5 kb, 1.5

kb, 2.0 kb, 2.3 kb, 4.0 kb, 6.5 kb, 9.5 kb and 20.0 kb. The polymerase chain reaction

(PCR)

using 3 difference primers ( t e a , catl and drfAl) to detect the resistance genes of tetracycline, chloramphenicol and trimetophrim- sulphametoxazol demontrated that 6 isolates from chickens harbouring tetracycline genes (tetA), one tetA gene was amplified fiom chromosome of S. Enteritidis h m egg, and only 1 chIoramphenico1 gen (catl) was amplified fiom chromosome of S. Enteritidis from human. -The trimetophrim-sulphametoxazol genes (drfAl) could not be detected in this study.

Key words: Salmonelia Enteritidis, 'intimicrobe, resistance genes, chicken, eggs, . .

(4)

ABSTRAK

ANNI _{KUSUMANINGSIH.} _{Profil dan} _{Gen Resistensi Antimikroba} Salmonella Enteritidis asal aysm, telur, dan manusia. Dibawah bimbingan Hj. MIRNAWATI SUDARWANTO,

I

W A Y A N

TEGW

WIBAWAN,

ROCHMAN NA'M and

SUPAR

Salmonellosis yang disebabkan oleh Salmonella Enteritidis merupakan salah satu penyakit penting yang banyak ditemukan pada industri peternakan. Infeksi S. Enteritidis pada ayam tidak meriimbullran gejala klinis, tetapi bakteri persisten dalam jaringan limfatik. Penularan ke maxlusia disebabkan oleh konsumsi pangan asal ayam, telur, dan olahannya yang tercemar dan d i i a k tidak sempurna. Tujuan penelitian

ini

untuk

membuat antisera somatik 0 (9) d m flagela H (m) untuk seroming S. Enteritidis; mendapatkan isolat S. Enteritidis asal ayam, telur,

dan

manusia; mengetahui profl resistensi antimikroba isolat S. Enteritidis; mengetahui jumlah d m ukuran plasmid S. Enteritidis, serta mengamplifikasi gen resistensi tetrasiklin, klodenikol, dan trimetoprim- sulfametoksasol dari plasmid

dan

kromosom S. Enteritidis. Sampel usapan rektal, pasangan hati dan jantung, serta telur ayam diambil dari peternakan dan pasar di Bogor, Sukaburni, Cianjur, dan Bandung. Sarnpel usapan rektdfeses manusia diambil dari -rumah sakit, dan isolat Salmonella dari Laboratorium Mikrobiologi, Fakultas Kedokteran Universitas Indonesia, Depok dan laboratoriurn swasta, Bogor. Sebanyak 33 isolat S. Enteritidis dari Baljtvet Culture Collection, Bdai Besar Penelitian Veteriner juga dipakai dalam penelitian ini. Dalam penelitian ini dapat diisolasi 1 1 isolat S. Enteritidis asal ayam, 14 isolat asal-telur, dan 13 isolat

a d manusia sebanYak 63 dari 68 (90.1%) isolat S. Enteritidis resisten terhadap streptomisin, neomisin, gentamisin, kloramfenikol, tetrasiklin, oksitetrasiklin, doksisiklin, trimetoprim-sulfametoksasol, siprofloksasin, dan enrofloksasin. Isolat S. Enteritidis asal ayam, telur, dan manusia telah resisten terhadap 2 sampai 9 jenis antimikroba, terbanyak multiresisten terhadap streptomisin, neomisin, tetrasiklin, doksisiklin, dan siprofloksasin. NiIai minimum inhibitory concenfration (MIC) dari keduapuluh l h isolat S. Enteritidis terhadap kloramfeniicol antam 12.5-50.0 pg/ml, tetrasiktin antara 0.8-25.0 pg/ml, trime~~rim-sulfametoksasol antara 6.3-50.0 pg/ml dan siprofloksasin <1,0 pg/ml. Hasi~ isolasi plasmid menunjukkan bahwa 2.1

dari

25 (84.0%) isolat S. Enteritidis asal ayam, telw; dan manusia membawa 1-6 pl&mid dengan ukuran 0.5

kt>,

1.5 kb, 2.0 kb, 2.3 kb, 4.6

kb,

6.5 kb, 9.5 kb, dan 20.0 kb, plasmid dengan ukuran 20.0 kb dibawa oleh semua isolat S. Enteritidis. Hasil a m p l i f h i gen resistensi menggunakan 3 pasang primer (tetA, cat 1, dan drfA I) untuk mengamplifikasi gen resistensi tetrasiklin, k l o d e n i k o l , dan trimetoprim-sulfametoksasol mecunjukkan bahwa gen resistensi tetrasiklin (MA) dapat diamplifikasi dari 6 plasmid S. Enteritidis asal ayam dan 1 kromosom S. Enteritidis asal telur, serta gen resistensi k l o d e n i k o l ( a t l ) dapat diamplifikasi dari

1 kromosom S.

Enteritidis asal rnanusia, sedarLgkan gen resistensi trimetoprim-sulfametoksasol belum dapat diamplifikasi baik dari plasmid maupun kromosom S. Enteritidis asal ayam, telur,

dan

manusia.

(5)

Hak cipta milik Institut Pertanian Bogor, tahun 2007

Hak cipta dilindungi

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PROPIL DAN GEN RESISTENSI ANTIMIKROBA

Salmonella

Enteritidis ASAL AYAM, TELUR,

DAN MANUSIA

ANN1 KUSUMANINGSIH

SEKOLAH PASCASARJANA

INSTITUT

. .

PERTANIAN

BOGOR

. .

BOGOR

.

(134)

ABSTRACT

ANNI

KUSUMANINGSIH.

The Profile and Genes of Antimicrobes Resistance of Safmoneffa Enteritidis from Chickens, Eggs, and Human. Supervised by Hj. MIRNAWATI SUDARWANTO, I WAYAN TEGUH WIBAWAN, ROCHMAN NA'IM and SUPAR

SalmoneIlosis associated with Salmonella Enteritidis infection is considered

as

an important disease in poultry industries. The disease in chicken sometimes shows no clinical signs, but the agents persist in the lymphatic tissues and contaminated their produck The infection in human due to consume chickens, eggs, and theirs products contaminated

with

S. Enteritidis. The objectives of this study were: to produce

0

somatic (9) and

H

flagella

(m)

antisera for serotyping; to obtain

S.

Enteritidis isolates from chickens, eggs, and human origin; to study the antimicrobial resistance profile of S. Enteritidis isolates; to observed the number and size of plasmid from S. Enteritidis; and to detect the resistance genes of tetracycline, chloramphenicol, and trimetoprim- sulphametoxazol from S. Enteritidis plasmids and chromosomes. Samples of chicken cloaca1 swabs, livers and hearts, and eggs were collected h r n _{farms and}

markets

in Bogor, Sukabumi, Cianjur, and Bandung; swab samples f b m human were also collected from hospital; some Salmonella spp. isolates were obtained fiom Laboratory of Microbiology, Medical School,

The

University of Indonesia, Jakarta; and private laboratory in Bogor. Thirty three isolates of S. Enteritidis fiom Balitvet Culture Colection werc: included in this study. Antimicrobe sensitivity assays of 68 S. Enteritidis indicated that 90.1% of these isolates were isolated from chickens, 14 isolates fiom eggs, and 13 isolates h m human. Sixty three of 68 (90,10%) S. Enteritidis isolates were resistance to streptomycin, neornycin, gentamycin, chloramphenicol, tetracycline, oxytetracycline, doxicycline, trimetophrim, ciproflocacin, and enrofloxacin-The multiresistance

between

2 to 9 antimicrobial agents: such as streptomycin, neomicyn, gentamicin tetrrrcyclin, doxicycline, and ciprofloxacine. Twenty five isolates of S. Enteritidis fiom chkken, eggs, and 'hLuriar1 tested qualitatively against chloramphenicol, tetracycline,

-tdmetophrim-sulhpmetoxazol,

and ciprofloxacin demonstrated MICs - of betwe"e6 12.50-50.0 pg/rnl, 0.8-50.0 pglml, 6.3-50.0 pg/ml, and less than 0.1 pgml respectively. Twenty one of 25 (84%) isolates which were resistant to 10 different antimicrobes carried 1-6 plasmids with differences in sizes: 0.5 kb, 1.5

kb, 2.0 kb, 2.3 kb, 4.0 kb, 6.5 kb, 9.5 kb and 20.0 kb. The polymerase chain reaction

(PCR)

using 3 difference primers ( t e a , catl and drfAl) to detect the resistance genes of tetracycline, chloramphenicol and trimetophrim- sulphametoxazol demontrated that 6 isolates from chickens harbouring tetracycline genes (tetA), one tetA gene was amplified fiom chromosome of S. Enteritidis h m egg, and only 1 chIoramphenico1 gen (catl) was amplified fiom chromosome of S. Enteritidis from human. -The trimetophrim-sulphametoxazol genes (drfAl) could not be detected in this study.

Key words: Salmonelia Enteritidis, 'intimicrobe, resistance genes, chicken, eggs, . .

(135)

ABSTRAK

ANNI _{KUSUMANINGSIH.} _{Profil dan} _{Gen Resistensi Antimikroba} Salmonella Enteritidis asal aysm, telur, dan manusia. Dibawah bimbingan Hj. MIRNAWATI SUDARWANTO,

I

W A Y A N

TEGW

WIBAWAN,

ROCHMAN NA'M and

SUPAR

Salmonellosis yang disebabkan oleh Salmonella Enteritidis merupakan salah satu penyakit penting yang banyak ditemukan pada industri peternakan. Infeksi S. Enteritidis pada ayam tidak meriimbullran gejala klinis, tetapi bakteri persisten dalam jaringan limfatik. Penularan ke maxlusia disebabkan oleh konsumsi pangan asal ayam, telur, dan olahannya yang tercemar dan d i i a k tidak sempurna. Tujuan penelitian

ini

untuk

membuat antisera somatik 0 (9) d m flagela H (m) untuk seroming S. Enteritidis; mendapatkan isolat S. Enteritidis asal ayam, telur,

dan

manusia; mengetahui profl resistensi antimikroba isolat S. Enteritidis; mengetahui jumlah d m ukuran plasmid S. Enteritidis, serta mengamplifikasi gen resistensi tetrasiklin, klodenikol, dan trimetoprim- sulfametoksasol dari plasmid

dan

kromosom S. Enteritidis. Sampel usapan rektal, pasangan hati dan jantung, serta telur ayam diambil dari peternakan dan pasar di Bogor, Sukaburni, Cianjur, dan Bandung. Sarnpel usapan rektdfeses manusia diambil dari -rumah sakit, dan isolat Salmonella dari Laboratorium Mikrobiologi, Fakultas Kedokteran Universitas Indonesia, Depok dan laboratoriurn swasta, Bogor. Sebanyak 33 isolat S. Enteritidis dari Baljtvet Culture Collection, Bdai Besar Penelitian Veteriner juga dipakai dalam penelitian ini. Dalam penelitian ini dapat diisolasi 1 1 isolat S. Enteritidis asal ayam, 14 isolat asal-telur, dan 13 isolat

a d manusia sebanYak 63 dari 68 (90.1%) isolat S. Enteritidis resisten terhadap streptomisin, neomisin, gentamisin, kloramfenikol, tetrasiklin, oksitetrasiklin, doksisiklin, trimetoprim-sulfametoksasol, siprofloksasin, dan enrofloksasin. Isolat S. Enteritidis asal ayam, telur, dan manusia telah resisten terhadap 2 sampai 9 jenis antimikroba, terbanyak multiresisten terhadap streptomisin, neomisin, tetrasiklin, doksisiklin, dan siprofloksasin. NiIai minimum inhibitory concenfration (MIC) dari keduapuluh l h isolat S. Enteritidis terhadap kloramfeniicol antam 12.5-50.0 pg/ml, tetrasiktin antara 0.8-25.0 pg/ml, trime~~rim-sulfametoksasol antara 6.3-50.0 pg/ml dan siprofloksasin <1,0 pg/ml. Hasi~ isolasi plasmid menunjukkan bahwa 2.1

dari

25 (84.0%) isolat S. Enteritidis asal ayam, telw; dan manusia membawa 1-6 pl&mid dengan ukuran 0.5

kt>,

1.5 kb, 2.0 kb, 2.3 kb, 4.6

kb,

6.5 kb, 9.5 kb, dan 20.0 kb, plasmid dengan ukuran 20.0 kb dibawa oleh semua isolat S. Enteritidis. Hasil a m p l i f h i gen resistensi menggunakan 3 pasang primer (tetA, cat 1, dan drfA I) untuk mengamplifikasi gen resistensi tetrasiklin, k l o d e n i k o l , dan trimetoprim-sulfametoksasol mecunjukkan bahwa gen resistensi tetrasiklin (MA) dapat diamplifikasi dari 6 plasmid S. Enteritidis asal ayam dan 1 kromosom S. Enteritidis asal telur, serta gen resistensi k l o d e n i k o l ( a t l ) dapat diamplifikasi dari

1 kromosom S.

Enteritidis asal rnanusia, sedarLgkan gen resistensi trimetoprim-sulfametoksasol belum dapat diamplifikasi baik dari plasmid maupun kromosom S. Enteritidis asal ayam, telur,

dan

manusia.

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