DETECTION OF SALMONELLA SPP. IN COMMERCIAL EGGS FROM LAYER CHICKEN FARMS AND TRADITIONAL MARKETS IN BALI

This study aims to early detect Salmonella spp. and identify serotypes in commercial chicken eggs from layer chicken farms and traditional markets in Bali. Early detection study of Salmonella spp. was carried out by conventional bacteriological methods, while serotype identification by duplex Polymerase Chain Reaction (d-PCR) test against the invA gene from Salmonella spp. and the sefA gene from Salmonella enteritidis. Egg samples in this study were taken from 10 layer chicken farms in Bali which included districts of Bangli, Gianyar, Tabanan and Karangasem. Egg samples from traditional markets were taken from 18 traditional markets from the districts of Bangli, Gianyar, Tabanan, Karangasem, Badung, and Denpasar City. Samples were eggshells and egg whites. Analysis of positive results from Salmonella spp. described descriptively. The results showed that eggshells and white eggs from all of the layer chicken farms are negative contaminated with Salmonella spp. (0%). In eggshell samples taken from the traditional markets of Taman Bali and Tulikup from the districts of Bangli and Gianyar, positive with Salmonella spp. (11,1%) by conventional bacteriological tests. In the duplex Polymerase Chain Reaction test, S. enteritidis serotypes were identified. The finding contamination of Salmonella enteritidis in commercial chicken eggs from traditional markets require periodically detection to prevent the occurrence of salmonellosis due to consumption of contaminated chicken eggs in traditional markets in Bali.

DUPLEX POLYMERASE CHAIN REACTION UNTUK DETEKSI SIMULTAN KOI HERPESVIRUS DAN Aeromonas hydrophila PADA IKAN MAS (Cyprinus carpio)

Koi herpesvirus (KHV) dan Aeromonas hydrophila adalah patogen yang dapat mengkoinfeksi ikan mas secara bersamaan. Penelitian ini bertujuan untuk mengembangkan metode duplex polymerase chain reaction (dPCR), deteksi simultan untuk diagnosis KHV dan bakteri Aeromonas hydrophila pada ikan mas. Dua pasang primer yang menargetkan sekuen spesifik SphI dan gen aerolisin, yang sering digunakan untuk mendeteksi KHV dan A. hydrophila dalam uji reaksi tunggal PCR dan menghasilkan target pita PCR 290 bp dan 417 bp. Hasil penelitian menunjukkan bahwa metode duplex PCR dapat mendeteksi ganda infeksi KHV dan A. hydrophila pada ikan mas dan metode ini lebih efektif mendeteksi dua patogen secara bersamaan dalam satu reaksi PCR pada suhu pradenaturasi, 94°C selama dua menit, denaturasi pada suhu 95°C selama satu menit, annealing pada suhu, 55°C selama satu menit, dan 72°C selama satu menit, dengan 30 siklus amplifikasi dan final extention pada suhu 72°C selama lima menit. Metode dPCR untuk deteksi simultan kedua patogen adalah salah satu metode yang dapat diaplikasikan untuk deteksi koinfeksi virus dan bakteri dalam satu reaksi PCR.Koi herpesvirus (KHV) and Aeromonas hydrophila are pathogens that can co-infect common carp. This study aimed to develop a duplex polymerase chain reaction (dPCR) method to detect KHV and Aeromonas hydrophila in common carp simultaneously. Two pairs of primers targeted the specific sequences of SphI and aerolysin genes, often used in detecting KHV and A. hydrophila, in a single PCR reaction test and produced target bands of PCR 290 bp and 417 bp. This proposed method was more effective in simultaneously detecting the two pathogens in one PCR reaction at pre-naturation temperature of 94°C for two minutes, denaturation at 95°C for one minute, annealing at temperature, 55°C for one minute, and 72°C for one minute, with 30 cycles of amplification and final extension at 72°C for five minutes. The findings showed that the duplex PCR method could be used to double detect KHV and A. hydrophila infection in common carp. The duplex PCR method for simultaneous detection of both pathogens is one method that can be applied for the detection of co-infection of viruses and bacteria in a PCR reaction.

Detection of genes encoding ompW and ctxA of Vibrio cholerae isolated from shrimp and shellfish at Kedonganan fish market, Bali-Indonesia

<p>Contamination of pathogenic bacteria in food can lead to the emergence of foodborne disease. One of foodborne disease which often occurs in some developing countries such as Africa, Southeast Asia, and Latin America is cholera which is caused by <em>Vibrio cholerae</em>. The disease is transmitted through beverages and food, especially contaminated seafood. <em>V. cholerae</em> has several virulence factors including the outer membrane protein W <em>(ompW)</em> and cholerae toxin <em>(ctx).</em>The<em> ompW</em> acts as a protective barrier and can also be used as a marker specific species of <em>V. cholerae</em> and cholerae toxin is an enterotoxin responsible for the incidence of diarrhea in a cholera outbreak produced by pathogenic <em>V. cholerae</em>. This study was an observational study to determine the level of contamination of <em>V. cholerae</em> by detecting the outer membrane protein W <em>(ompW)</em> and cholerae toxin subunit A <em>(ctxA)</em> gene of <em>V. cholerae</em> in shrimp and shellfish sold at Kedonganan fish market. Samples were taken using total sampling technique and obtained 24 samples consisting of 14 shrimp samples and 10 shellfish samples. Samples were examined using culture methods and biochemical tests, and then further tested using Duplex Polymerase Chain Reaction (dPCR) to detect <em>ompW</em> and <em>ctxA</em> gene. The dPCR assay results showed 8 out of 14 (57.1%) samples from shrimp and 1 out of 10 (10%) samples from the shellfish positive carried <em>ompW</em> gene, and found no positive samples carrying the <em>ctxA</em> gene in samples derived from shrimp and shellfish. Chi square test analysis results indicated contamination of <em>V. cholerae</em> in shrimp was higher than shellfish based on <em>ompW</em> gene (p&lt;0.05). It can be concluded that the shrimp and shellfish at Kedonganan fish market are contaminated by <em>V. cholerae</em>. Further research is needed to detect the virulence factors besides <em>ompW</em> and <em>ctxA </em>of<em> V. cholerae</em> in seafood.</p><pre><strong> </strong></pre>Keywords: Foodborne disease, <em>Vibrio cholerae</em>, <em>ompW</em> gene<em>,</em> <em>ctxA</em> gene, and Duplex Polymerase Chain Reaction (dPCR).

Molecular characterization of Candida dubliniensis and Candida albicans in the oral cavity of drug abusers using duplex polymerase chain reaction

Background and Purpose: Candida dubliniensis is closely related to the most pathogenic and prevalent yeast, namely C. albicans. Candida species can opportunistically overgrow in vulnerable individuals and cause a variety of diseases. The current study aimed to identify and isolate C. dubliniensis species present in the Candida albicans species complex identified in the oral cavity of drug abusers. Materials and Methods: This study was conducted on 53 strains of C. albicans species complex, isolated from the oral mucosa of drug abusers in Isfahan, Iran. DNA extraction was accomplished through boiling procedure. Duplex polymerase chain reaction (PCR) was performed to amplify ITS1-5.8S-ITS2 region using four specific primers. Fungal species were identified based on the difference in the size of the bands created in the Agarose gel. Results: Out of the 53 isolates under study, 30 (56.6%) and 14 (26.4%) samples were identified as C. albicans and C. dubliniensis, respectively. In the remaining 9 samples (17%), both types of Candida species were confirmed. Conclusion: The findings of the present study revealed the presence of a noticeable amount of C. dubliniensis in the oral cavity of drug abusers. Therefore, the probable presence of this fungus should be considered during the examination of oral infection among this group. To date, no research has directly investigated this issue in Iran.