No abstract

At present, only a single Rhizopus species, R. microsporus, can be found in fresh tempeh produced in Java, Indonesia. The loss of diversity of Rhizopus in tempeh has been associated with the widespread use of commercial tempeh starter in Indonesia since the 2000s. However, the identities of the previous Rhizopus strains associated with tempeh, which have been preserved in a culture collection in Indonesia, have not been verified. The present study aimed to verify the identities of 22 Rhizopus strains isolated from tempeh produced using the traditional tempeh starters from the 1960s to the 2000s. Phylogenetic analysis based on the ITS regions in the rRNA gene sequence data, revealed that the Rhizopus strains belonged to the species R. arrhizus (five strains); R. delemar (14 strains); and R. microsporus (three strains). Verification of the identities of these Rhizopus strains in the present study confirmed the loss of diversity of Rhizopus species in tempeh produced in Indonesia, particularly in Java. Our findings confirmed that the morphological changes in Rhizopus species isolated from tempeh as a result of centuries of domestication.

Diagnosis of Neisseria gonorrhoeae infection is needed for patient therapy and for reducing this bacterial transmission in the population. The culture method is a gold standard method for N. gonorrhoeae detection, however it has low sensitivity. Among molecular methods with high sensitivity and specificity, SYBR Green real-time PCR is the potential method for N. gonorrhoeae detection. In this study, we developed an SYBR Green real-time PCR-based system assay for N. gonorrhoeae detection. Several PCR conditions were optimized and analyzed including primer annealing temperature, DNA template volume, the limit of detection (LoD), cross-reaction with others (bacteria, viruses, fungus, protozoa), and quality assurance. The results showed that the annealing temperature and DNA template volume were 60oC and 5 µL, respectively. The LoD was 29 DNA copies corresponding to 3 bacterial cells per reaction. No cross-reaction was detected for other bacteria, viruses, fungus and protozoa. The external quality assurances enrolled in 2019 and 2021 showed 100% concordance. The preliminary testing for clinical samples was also 100% concordance. In conclusion, the SYBR Green real-time PCR-based system assay developed in this study is promising for application in clinical laboratories.