SummaryThe herbicide 2,4,5‐trichlorophenoxyacetic acid (2,4,5‐T) was a major component of Agent Orange, which was used as a defoliant in the Vietnam War. Little is known about its degradation under anoxic conditions. Established enrichment cultures using soil from an Agent Orange bioremediation plant in southern Vietnam with pyruvate as potential electron donor and carbon source were shown to degrade 2,4,5‐T via ether cleavage to 2,4,5‐trichlorophenol (2,4,5‐TCP), which was further dechlorinated to 3,4‐dichlorophenol. Pyruvate was initially fermented to hydrogen, acetate and propionate. Hydrogen was then used as the direct electron donor for ether cleavage of 2,4,5‐T and subsequent dechlorination of 2,4,5‐TCP. 16S rRNA gene amplicon sequencing indicated the presence of bacteria and archaea mainly belonging to the Firmicutes, Bacteroidetes, Spirochaetes, Chloroflexi and Euryarchaeota. Desulfitobacterium hafniense was identified as the dechlorinating bacterium. Metaproteomics of the enrichment culture indicated higher protein abundances of 60 protein groups in the presence of 2,4,5‐T. A reductive dehalogenase related to RdhA3 of D. hafniense showed the highest fold change, supporting its function in reductive dehalogenation of 2,4,5‐TCP. Despite an ether‐cleaving enzyme not being detected, the inhibition of ether cleavage but not of dechlorination, by 2‐bromoethane sulphonate, suggested that the two reactions are catalysed by different organisms.

BK polyomavirus (BKV) is a common opportunistic pathogen in the community, infecting humans in their early life stages, often remaining in a latent state. BK virus-associated nephropathy occurs in 2 to 10% of kidney recipients, and up to 40 to 90% of these patients have permanent loss of graft function due to delay in diagnosis. In the present study, we have successfully optimized the component and thermal cycling for the long-PCR assay to amplify the BKV genome, including middle-concentration samples such as plasma (concentration about 105 copy/ml). We have successfully amplified the BKV genome in 41/50 samples, including 04 plasma and 39 urine samples. The optimized amplification protocol of full-length BKV genome includes the following steps: 1) Extraction of DNA-BKV from plasma and urine samples and quantification of BKV-DNA concentrations to ensure that about 105 copies/ml (plasma) and 107 copies/ml (urine); 2) Each full-length genome reaction in the volume of 50 μl containing 1X Phusion HF master mix (Thermo Scientific, USA); 0.5 μM of wBK_a(F+R) primers, 3-5 μl of genomic DNA. Cycling conditions were 98°C for 30 s, followed by 35 cycles of 98°C for 10 s; 58°C for 20 s; 72°C for 2 min, and 72ºC for 6 min and a 4oC hold; 3) All 5.1 kb purified PCR were verified and confirmed by restriction enzymes XbaI, EcoRI, and direct sequencing.

Nitrogen removal with biological methods plays a crucial role in wastewater treatment technology. The treatment begins with the oxidation of ammonia to nitrite to facilitate the subsequent nitrification and denitrification. Various strains of ammonia-oxidising bacteria have been reported. In this study, we use three Bacillus bacteria isolated from swine wastewater to oxidise ammonia. Different initial densities (103, 104, 105, and 106 CFU·mL–1) of each strain were examined. The results show that the combination of all the bacteria at a ratio of 1:1:1 and a density of 105 CFU·mL–1 exhibits the most effect. The findings contribute to the diversity of ammonia-oxidising bacterial species and pose a great potential for applying these strains in wastewater treatment.

Ebola virus is a deadly causative agent with a high mortality rate of up to 90%, therefore it has been classified by the Center for Disease Control and Prevention (CDC) as a category A biological agent. The World Health Organization (WHO) recommended using RT-PCR based assays to rapidly detect the virus. In the present study, we established an in-house assay for detection of Zaire ebolavirus via real-time RT-PCR. The nucleotide sequence of the Zaire ebolavirus nucleoprotein (NP) gene was retrieved from the Genbank for designing primer pairs and probes using Primer Express 3.0 software. The RNA positive control was generated by in vitro RNA transcript synthesis. The optimal components in the 20 μl final volume of the real-time RT-PCR assay were 10 μl 2X QuantiTectProbeRT-PCR master mix, 0,6 μM of each primer, 0,1 μM of the probe, 0,2 μl RT mix and 5 μl of RNA template. The thermal cycle conditions were as follows: 50oC for 30 min, 95°C for 15 min, then 45 cycles of 15 s at 94°C, 60s at 60°C. The limit of detection of the assay was 100 copies/reaction and 1414 FFU/ml with the positive RNA panel and sample panel of RNA extracted from cell culture supernatants of cells infected with Zaire ebolavirus 2014/Gueckedou-C05, respectively. The specificity of this assay was 100% when tested with the positive RNA panel of Ebola virus and other haemorrhagic fever viruses. In conclusion, we successfully established an in-house real-time RT-PCR assay for detection of Zaire ebolavirus in Vietnam with a limit of detection of 1414 FFU/ml and specificity of 100%.

In Vietnam, nasopharyngeal carcinoma (NPC) is the eighth most common cause of death from cancer. Cell-free Epstein Barr virus DNA (cf-EBV DNA) was reported to be present in almost all NPC patients. However, currently available assays in Vietnam can detect cf-EBV DNA in only 67.6% of NPC patients, thus leaving 32.4% of cancer cases undetected. Therefore, in this study, we aim to develop a highly sensitive quantitative PCR (qPCR) assay that measures the load of cf-EBV DNA for the purpose of early detection of NPC, and then evaluate the sensitivity and the specificity of the developed qPCR assay on the clinical samples. The major methods used in this study include primer/TaqMan probe design, cf-DNA extraction, optimization of qPCR assay and statistical analysis. Using an international standard panel from the Chinese University of HongKong, the linear range of developed qPCR assay is from 50-150,000 copies/ml (R2 = 0.99613) and the detection limit has been shown to be 25 copies/ml. The developed assay could detect cf-EBV DNA with a sensitivity of 96.9% (31/32 NPC patients) and cf-EBV DNA has not been detected in 103 out of 105 healthy controls, which corresponds to a specificity of 98%. Consequently, the performance of the optimal assay has achieved remarkably high sensitivity and specificity. Moreover, the detection limit of our optimal qPCR assay is 25 copies/ml of plasma, which is at least ten times better than other assays tested in recent studies in Vietnam. This developed qPCR assay will also form the basis for further studies in Vietnam and will open many new applications in management of NPC.