Water and food are major environmental transmission routes for Cryptosporidium, but our ability to identify the spectrum of oocyst contributions in current performance-based methods is limited. Determining risks in water and foodstuffs, and the importance of zoonotic transmission, requires the use of molecular methods, which add value to performance-based morphologic methods. Multi-locus approaches increase the accuracy of identification, as many signatures detected in water originate from species/genotypes that are not infectious to humans. Method optimisation is necessary for detecting small numbers of oocysts in environmental samples consistently, and further work is required to (i) optimise IMS recovery efficiency, (ii) quality assure performance-based methods, (iii) maximise DNA extraction and purification, (iv) adopt standardised and validated loci and primers, (v) determine the species and subspecies range in samples containing mixtures, and standardising storage and transport matrices for validating genetic loci, primer sets and DNA sequences.

The purpose of this study was to investigate the impact of microaeration on the fermentation process during anaerobic co-digestion of brown water (BW) and food waste (FW). This was achieved by daily monitoring of reactor performance and the determination of its bacterial consortium towards the end of the study. Molecular cloning and sequencing results revealed that bacteria within phyla Firmicutes and Bacteriodetes represented the dominant phylogenetic group. As compared to anaerobic conditions, the fermentation of BW and FW under microaeration conditions gave rise to a significantly more diverse bacterial population and higher proportion of bacterial clones affiliated to the phylum Firmicutes. The acidogenic reactor was therefore able to metabolize a greater variety of substrates leading to higher hydrolysis rates as compared to the anaerobic reactor. Other than enhanced fermentation, microaeration also led to a shift in fermentation production pattern where acetic acid was metabolized for the synthesis of butyric acid.

Cyclospora cayetanensis is a coccidian parasite that causes diarrheal illness outbreaks worldwide. The development of new laboratory methods for detection of C. cayetanensis is of critical importance because of the high potential for environmental samples to be contaminated with a myriad of microorganisms, adversely impacting the specificity when testing samples from various sources using a single molecular assay. In this study, a new sequencing-based method was designed targeting a specific fragment of C. cayetanensis cytochrome oxidase gene and developed as a complementary method to the TaqMan qPCR present in the U.S. FDA BAM Chapter 19b and Chapter 19c. The comparative results between the new PCR protocol and the qPCR for detection of C. cayetanensis in food and water samples provided similar results in both matrices with the same seeding level. The target region and primers in the protocol discussed in this study contain sufficient Cyclospora-specific sequence fidelity as observed by sequence comparison with other Eimeriidae species. The sequence of the PCR product appears to represent a robust target for identifying C. cayetanensis on samples from different sources. Such a sensitive method for detection of C. cayetanensis would add to the target repertoire of qPCR-based screening strategies for food and water samples.