METHODS AND RESULTS: The Rhizomucor pusillus proteinase (RPP) gene was sub-cloned into a pALF expression vector. The recombinant pALF-RPP vector was then electro-transferred into Lactococcus lactis. Finally, the milk coagulation ability of recombinant L. lactis carrying a RPP gene was evaluated. Nucleotide sequencing of DNA insertion from the clone revealed that the RPP activity corresponded to an open reading frame consisting of 1218 bp coding for a 43·45 kDa RPP protein. The RPP protein assay results indicated that the highest RPP enzyme expression with 870 Soxhlet units (SU) per ml and 7914 SU/OD were obtained for cultures which were incubated at pH 5·5 and 30°C. Interestingly, milk coagulation was observed after 205 min of inoculating milk with recombinant L. lactis carrying the RPP gene.

CONCLUSION: The recombinant L. lactis carrying RPP gene has the ability to function as a starter culture for acidifying and subsequently coagulating milk by producing RPP as a milk coagulant agent.

METHODS AND RESULTS: To explore the dynamics of microbial population in mushroom substrate during commercial mushroom cultivation and how microbiota might play a role in green-mould contamination, we applied both culturing and targeted metagenomics approaches to identify microbiota in noncomposted sawdust substrates at different cultivation stages. The microbiological analysis showed that the green-mould contaminated substrates harboured higher total mesophilic bacteria count. The green-moulds isolated from the contaminated mushroom substrates were identified as Trichoderma pleurotum (n = 15; 93.8%) and Graphium penicillioides (n = 1; 6.3%). To our surprise, the targeted metagenomic analysis revealed that Graphium comprised 56.3% while Trichoderma consisted of only 36.1% of the total fungi population, suggesting that green-mould contamination might not be caused by Trichoderma alone, but also Graphium that grows very slowly in the laboratory.

CONCLUSION: It is worthwhile to note that G. penicillioides was also isolated in the early stages of mushroom cultivation, but not T. pleurotum. The results indicated that the structure and composition of the bacterial population in the mushroom substrate varied and the bacterial population shifted along the cultivation process.

METHODS AND RESULTS: The pulp of red pitahaya and the leaves of red spinach were extracted using methanol followed by subfractionation to obtain betacyanin fraction. The anti-biofilm activity was examined using broth microdilution assay on polystyrene surfaces and expressed as minimum biofilm inhibitory concentration (MBIC). The betacyanin fraction from red spinach showed better anti-biofilm activity (MBIC: 0·313-1·25 mg ml-1 ) against five Staph. aureus strains while the betacyanin fraction from red pitahaya showed better anti-biofilm activity (MBIC: 0·313-0·625 mg ml-1 ) against four P. aeruginosa strains. Both betacyanin fraction significantly reduced hydrophobicity of Staph. aureus and P. aeruginosa strains. Numbers of Staph. aureus and P. aeruginosa attached to polystyrene were also reduced without affecting their cell viability.

CONCLUSION: Betacyanins can act as anti-biofilm agents against the initial step of biofilm formation, particularly on a hydrophobic surface like polystyrene.

METHODS AND RESULTS: The discussion ranged from examining scientific literature supporting the efficacy of established prebiotics, to the prospects for establishing health benefits associated with novel compounds, isolated from different sources.

CONCLUSIONS: While many promising candidate prebiotics from across the globe have been highlighted in preliminary research, there are a limited number with both demonstrated mechanism of action and defined health benefits as required to meet the prebiotic definition. Prebiotics are part of a food industry with increasing market sales, yet there are great disparities in regulations in different countries. Identification and commercialization of new prebiotics with unique health benefits means that regulation must improve and remain up-to-date so as not to risk stifling research with potential health benefits for humans and other animals.

METHODS AND RESULTS: Abnormal behaviour, clinical signs, postinjection survival and histopathology (kidney, liver, eye and brain) were measured. Cumulative mortality of CON+ , free cells, ALG and treatments (F1-F7) was 30, 24, 22, 19, 17, 17, 16, 14, 14 and 12 out of 30 fish and the survival rates for E. faecium ABRIINW.N7 microencapsulated in an alginate-BS blend with 0·5, 1, 1·5, 2, 2·5 and 3% fenugreek were 43, 43, 47, 53, 53 and 60%, respectively. After the incorporation of fenugreek with the alginate-BS blend, there was an 8-21% increase in probiotic cell viability. Furthermore, the survival rate for the alginate-BS blend with 2·5 and 3% fenugreek (F6 and F7) was significantly (P ≤ 0·05) higher than other blends. The highest encapsulation efficiency, viability in gastrointestinal conditions and during storage time and excellent antipathogenicity against S. iniae were observed in alginate-BS +3% fenugreek formulation (F7).

CONCLUSIONS: It is recommended that probiotic strains like E. faecium ABRIINW.N7 in combination with local herbal gums, such as BS and fenugreek plus alginate, can be used as a suitable scaffold and an ideal matrix for the encapsulation of probiotics.