Development and validation of multiplex real-time PCR for simultaneous detection of six bacterial pathogens causing lower respiratory tract infections and antimicrobial resistance genes.

BACKGROUND: Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Escherichia coli, Streptococcus pneumoniae and Staphylococcus aureus are major bacterial causes of lower respiratory tract infections (LRTIs) globally, leading to substantial morbidity and mortality. The rapid increase of antimicrobial resistance (AMR) in these pathogens poses significant challenges for their effective antibiotic therapy. In low-resourced settings, patients with LRTIs are prescribed antibiotics empirically while awaiting several days for culture results. Rapid pathogen and AMR gene detection could prompt optimal antibiotic use and improve outcomes. METHODS: Here, we developed multiplex quantitative real-time PCR using EvaGreen dye and melting curve analysis to rapidly identify six major pathogens and fourteen AMR genes directly from respiratory samples. The reproducibility, linearity, limit of detection (LOD) of real-time PCR assays for pathogen detection were evaluated using DNA control mixes and spiked tracheal aspirate. The performance of RT-PCR assays was subsequently compared with the gold standard, conventional culture on 50 tracheal aspirate and sputum specimens of ICU patients. RESULTS: The sensitivity of RT-PCR assays was 100% for K. pneumoniae, A. baumannii, P. aeruginosa, E. coli and 63.6% for S. aureus and the specificity ranged from 87.5% to 97.6%. The kappa correlation values of all pathogens between the two methods varied from 0.63 to 0.95. The limit of detection of target bacteria was 1600 CFU/ml. The quantitative results from the PCR assays demonstrated 100% concordance with quantitative culture of tracheal aspirates. Compared to culture, PCR assays exhibited higher sensitivity in detecting mixed infections and S. pneumoniae. There was a high level of concordance between the detection of AMR gene and AMR phenotype in single infections. CONCLUSIONS: Our multiplex quantitative RT-PCR assays are fast and simple, but sensitive and specific in detecting six bacterial pathogens of LRTIs and their antimicrobial resistance genes and should be further evaluated for clinical utility.

The clinical features and genomic epidemiology of carbapenem-resistant Acinetobacter baumannii infections at a tertiary hospital in Vietnam.

OBJECTIVES: To characterise the clinical features of Acinetobacter baumannii infections and investigate the phylogenetic structure and transmission dynamics of A. baumannii in Vietnam. METHODS: Between 2019 and 2020, a surveillance of A. baumannii (AB) infections was conducted at a tertiary hospital in Ho Chi Minh City, Vietnam. Risk factors for in-hospital mortality were analysed using logistic regressions. Whole-genome sequence data were used to characterise genomic species, sequence types (STs), antimicrobial resistance genes, surface antigens, and phylogenetic relatedness of AB isolates. RESULTS: Eighty-four patients with AB infections were enrolled in the study, 96% of whom were hospital-acquired. Half of the AB isolates were identified from ICU-admitted patients, while the remaining isolates were from non-ICU patients. The overall in-hospital mortality was 56%, with associated risk factors including advanced age, ICU stay, exposure to mechanical ventilation/central venous catheterization, pneumonia as source of AB infection, prior use of linezolid/aminoglycosides, and AB treatment with colistin-based therapy. Nearly 91% of isolates were carbapenem-resistant; 92% were multidrug-resistant; and 6% were colistin-resistant. ST2, ST571, and ST16 were the three dominant carbapenem-resistant A. baumannii (CRAB) genotypes, exhibiting distinct AMR gene profiles. Phylogenetic analysis of CRAB ST2 isolates together with previously published ST2 collection provided evidence of intra- and inter-hospital transmission of this clone. CONCLUSIONS: Our study highlights a high prevalence of carbapenem resistance and multidrug resistance in A. baumannii and elucidates the spread of CRAB within and between hospitals. Strengthening infection control measures and routine genomic surveillance are crucial to reducing the spread of CRAB and detecting novel pan-drug-resistant variants in a timely fashion.

Vertical distributions and risk assessment of phosphorus in core sediments from the Can Gio coastal district in southern Vietnam.

In this study, total phosphorus (P) and P released forms were measured in core sediments from the areas affected by human settlement and shrimp farming activities and the core zone of the Can Gio Biosphere Reserve, a coastal district in south Vietnam. Furthermore, ecological risk assessment and parameters controlling P release from sediments were investigated, including pH, major elements (Al-Ca-Mg-Fe), and fine fraction. The average total amount of P in the sediments varied from 287 to 669 mg/kg, with significantly lower values being observed in the mangrove biosphere reserve area. According to the results of the correlation analysis, organic matter was the primary source of P in the sediments, but the majority of the P released was inorganic. Positive correlations were found between Fe and non-apatite inorganic P (NAIP) and apatite P (AP), as well as intercorrelations between P fractions (r = 0.40-0.79, p < 0.05), suggesting that Fe might be the controlling factor of P release in the investigated sediments. The rank orders of concentrations of P forms were significantly different between the areas. The range of P forms was AP (35-248 mg/kg) > NAIP (63-201 mg/kg) > LP (labile P) (4-25 mg/kg) in the human settlement and aquaculture areas and NAIP (84-99 mg/kg) > AP (20-38 mg/kg) > LP (7-12 mg/kg) in the mangrove biosphere reserve area. Risk assessment based on the total concentration of P and the availability of P from a single extraction suggested a relatively low risk of P from sediment as an internal load in the studied areas.

Survey of knowledge, attitude and practice of healthcare professionals on dengue transmission, diagnosis and clinical classification.

BACKGROUND: To investigate the knowledge, attitudes, and practices of the healthcare professionals (HCPs) including physicians and nurses regarding dengue transmission, diagnosis and clinical classification using the warning signs of World Health Organization (WHO) 2009 classification. RESULTS: Out of 471 respondents from three countries, 80.9% of physicians and 74% of nurses did not receive previous training regarding the dengue infection. The majority of respondents could identify the primary dengue vector (86%), while only a third of HCPs knew the biting time of dengue mosquitoes. Only half of our respondents knew about immunity induced by serotypes; Moreover, half of our participants could determine the diagnostic tests. On the other hand, about 90% of the respondents took responsibility for talking to the patients about preventive measures. Our respondents also showed wide variations in definition of warning signs listed in the WHO 2009 classification. Multivariate analysis linked the impact of different cofactors including prior training on dengue infection, type of profession, frequency of taking care of dengue patients and country on how HCPs defined these warning signs. CONCLUSIONS: This study could declare the variation in employing the warning signs listed in the WHO 2009 classification. We have figured that most of the HCPs did not take prior training on the dengue viral infection; Also, we found gaps in the knowledge regarding various topics in dengue fever. This paper recommends the gathering of efforts to establish the proper knowledge of dengue infection and the warning signs listed by the WHO.

Yeast cell surface displaying VP28 antigen and its potential application for shrimp farming.

VP28 is an envelope protein of White Spot Syndrome Virus (WSSV), which has been shown in previous studies to induce a high immune response in shrimp. VP28 has been produced in some host systems such as Escherichia coli, Bacillus subtilis, and Pichia pastoris as free protein. Here we showed a new strategy of anchoring VP28 on the Saccharomyces cerevisiae yeast surface and using the yeast cell extract combined with probiotic as an oral vaccine for shrimp farming. We have successfully constructed a recombinant yeast cell capable of expressing VP28 on the cell surface. The feeding diet combined with VP28 anchored yeast cell extract provided significant assurance to Litopenaeus vannamei, challenged by WSSV, resulting in a relative percent survival (RPS) of 87.10 ± 2.15%. Interestingly, the utilization of VP28 anchored yeast cell extract could enhance the efficiency of probiotic strains like Lactobacillus and Bacillus on shrimp farming. The results in both laboratory scales and field trials using extract of VP28 displaying Saccharomyces showed a growth-promoting effect in shrimp, assessed through average shrimp weight. Taken together, our results in this study demonstrated a new successful strategy of using yeast cell surface as a tool to produce VP28-based oral vaccine for shrimp aquaculture. KEY POINTS: • A new strategy of using VP28 antigen as anchored protein on S. cerevisiae yeast cell surface (S. cerevisiae::VP28) • The utilization of VP28 antigen and yeast as S. cerevisiae::VP28 extract enhanced potential protection of Litopenaeus vannamei against White Spot Syndrome Virus (RPS 87.10%) • The use of S. cerevisiae::VP28 extract increased efficiency of probiotic on shrimp growth-promoting effect either lab-scale or field trial.

Late Pleistocene-Holocene sedimentary evolution in the coastal zone of the Red River Delta.

The Red River Delta is considered one of the largest megadelta systems in Asia. The formation of this delta has been controlled by the continent-ocean interaction and sea-level fluctuation during the Cenozoic. In this study, we present a new sequence stratigraphic framework of the Red River Delta based on borehole lithofacies analysis and high resolution seismic data. The Late Pleistocene-Holocene sediments in the coastal zone of the Red River Delta were subdivided into three systems tracts: (1) the lowstand systems tract (LST) is characterized by a Late Pleistocene alluvial silty sand facies complex (arLSTQ1 3b); (2) the transgressive systems tract (TST) is illustrated by the coastal marsh facies complex and the lagoonal greenish-gray clay facies of Early-Middle Holocene (amt, mtTSTQ2 1-2); and (3) the highstand systems tract (HST) is composed of the Middle-Late Holocene deltaic clayish silt facies complex (amhHSTQ2 2-3). The boundaries between these three systems tracts are not isochronous, namely: (1) The LST-HST boundary has been associated with the Würm 2 Glaciation, which occurred at ~40-18 Ka.; (2) The TST-LST boundary is identified by a transgressive erosion surface, whose age ranges from ~12-5 Ka.; and (3) the HST-TST boundary is an unconformity between the submarine deltaic facies complex and the Middle Holocene marine flooding plain.

Pig Exposure and Health Outcomes in Hospitalized Infectious Disease Patients in Vietnam.

Many infectious diseases have a zoonotic origin, and several have had major public health implications. Contact with animals is a known risk factor for zoonotic infections, although there are limited data on disease symptoms and pathogens associated with contact with different animal species. The rise in pig production in Southeast Asia has contributed to the emergence and re-emergence of zoonotic infections caused by contact with pigs and pig products. To compare the symptom and pathogen profiles of hospitalized patients with and without pig contact, we collected data on disease symptoms, infecting pathogens, and animal contact behaviour from patients attending six hospitals across Vietnam between 2012 and 2016. Patients who had previous contact with pigs were more likely to have enteric disease than respiratory or central nervous system infections and were more likely to grow Escherichia coli and Shigella from stool culture than those without pig contact. Patients with enteric infections who kept pigs were also more likely to have a disease of unknown origin. Public health initiatives that account for differences in animal contact behaviours and offer more comprehensive diagnostics in high-risk individuals are needed if emergence and re-emergence of zoonotic disease is to be monitored and prevented.

PAS1-modified optical SIS sensor for highly sensitive and specific detection of toluene.

We report on a novel solution immersed silicon (SIS) sensor modified with bio-receptor to detect toluene. To perform this approach, bio-receptor PAS1 which specifically interacts with toluene was chosen as a capture agent for SIS ellipsometric sensing. We constructed wild PAS1 and mutant PAS1 (F46A and F79Y) which are toluene binding-defective. Especially, we utilized an easily accessible capturing approach based on silica binding peptide (SBP) for direct immobilization of PAS1 on the SiO2 surfaces. After the immobilization of SBP-tagged PAS1 to the sensing layers, PAS1-based SIS sensor was evaluated for its ability to recognize toluene. As a result, a significant up-shift in Psi (Ψ) was clearly observed with a low limit of detection (LOD) of 0.1 µM, when treated with toluene on wild PAS1-surface, but not on mutant PAS1-sensing layers, indicating the selective interactions between PAS1 and toluene molecule. The PAS1-SIS sensor showed no changes in Psi (Ψ), if any, negligible, when exposed to benzene, phenol, xylene and 4-nitrophenol as negative controls, thereby demonstrating the specificity of interaction between PAS1 and toluene. Taken together, our results strongly indicate that PAS1-modified ellipsometry sensor can provide a high fidelity system for the accurate and selective detection of toluene.

Efficient click chemistry towards novel 1H-1,2,3-triazole-tethered 4H-chromene-d-glucose conjugates: Design, synthesis and evaluation of in vitro antibacterial, MRSA and antifungal activities.

The series of 2-amino-7-propargyloxy-4H-chromene-3-carbonitriles 5a-t were synthesized from corresponding 2-amino-7-phydroxy-4H-chromene-3-carbonitriles 4a-t and propargyl bromide. Two procedures were used in these syntheses: K2CO3/acetone and NaH/DMF procedures with yields of 65-89% and 80-96%, respectively. 1H-1,2,3-Triazole-tethered 4H-chromene-d-glucose conjugates 7a-t were synthesized using click chemistry of propargyl ethers 5a-t and tetra-O-acetyl-ß-d-glucopyranosyl azide. Cu@MOF-5 was the optimal catalyst for this chemistry. The yields of 1H-1,2,3-triazoles were 80-97.8%. All triazoles 7a-t were evaluated in vitro for anti-microorganism activities. Among tested compounds with MIC values of 1.56-6.25 µM, there were four compounds against B. subtilis, four compounds against S. aureus, and four compounds against S. epidermidis; five compounds against E. coli, four compounds against K. pneumoniae, five compounds against P. aeruginosa, and six compounds against S. typhimurium. Compounds 7c,7d,7f,7h, and 7r had MIC values of 1.56-6.25 µM for three clinical MRSA isolates. Some compounds had inhibitory activities against four fungi, including A. niger, A. flavus, C. albicans, and S. cerevisiae, with MIC values of 1.56-6.25 µM. Some 1H-1,2,3-triazoles had comparatively low toxicity against RAW 264.7 cells.

Contamination status and potential release of trace metals in a mangrove forest sediment in Ho Chi Minh City, Vietnam.

Can Gio district is located in the coastal area of Ho Chi Minh City, southern Vietnam. Discharge of wastewater from Ho Chi Minh City and neighboring provinces to the rivers of Can Gio has led to concerns about the accumulation of trace metals (As, Cu, Cr, Ni, Pb, and Zn) in the coastal sediments. The main objective of this study was to assess the distribution of As, Cu, Cr, Ni, Pb, and Zn in surface and core sediments and to evaluate the contamination status in relation to local background values, as well as the potential release of these selected trace metals from sediments to the water environment. Sediment characteristization, including determination of fine fraction, pH, organic matter, and major elements (Al, Fe, Ca, K, Mg, and S), was carried out to investigate which parameters affect the trace metal enrichment. Fine fraction and Al contents were found to be the controlling proxies affecting the distribution of trace metals while other sediment characteristics did not show any clear influence on trace metals' distribution. Although As concentrations in the sediments were much higher compared to its reference value in other areas, the enrichment factor based on local background values suggests minor contamination of this element as well as for Cr, Cu, and Pb. Risk assessment suggested a medium to very high risk of Mn, Zn, and Ni under acidification. Of importance is also that trace metals in sediments were not easily mobilized by organic complexation based on their low extractabilities by ammonium-EDTA extraction.

Applications and Advances in Bioelectronic Noses for Odour Sensing.

A bioelectronic nose, an intelligent chemical sensor array system coupled with bio-receptors to identify gases and vapours, resembles mammalian olfaction by which many vertebrates can sniff out volatile organic compounds (VOCs) sensitively and specifically even at very low concentrations. Olfaction is undertaken by the olfactory system, which detects odorants that are inhaled through the nose where they come into contact with the olfactory epithelium containing olfactory receptors (ORs). Because of its ability to mimic biological olfaction, a bio-inspired electronic nose has been used to detect a variety of important compounds in complex environments. Recently, biosensor systems have been introduced that combine nanoelectronic technology and olfactory receptors themselves as a source of capturing elements for biosensing. In this article, we will present the latest advances in bioelectronic nose technology mimicking the olfactory system, including biological recognition elements, emerging detection systems, production and immobilization of sensing elements on sensor surface, and applications of bioelectronic noses. Furthermore, current research trends and future challenges in this field will be discussed.

Role of Environmental Factors in Shaping Spatial Distribution of Salmonella enterica Serovar Typhi, Fiji.

Fiji recently experienced a sharp increase in reported typhoid fever cases. To investigate geographic distribution and environmental risk factors associated with Salmonella enterica serovar Typhi infection, we conducted a cross-sectional cluster survey with associated serologic testing for Vi capsular antigen-specific antibodies (a marker for exposure to Salmonella Typhi in Fiji in 2013. Hotspots with high seroprevalence of Vi-specific antibodies were identified in northeastern mainland Fiji. Risk for Vi seropositivity increased with increased annual rainfall (odds ratio [OR] 1.26/quintile increase, 95% CI 1.12-1.42), and decreased with increased distance from major rivers and creeks (OR 0.89/km increase, 95% CI 0.80-0.99) and distance to modeled flood-risk areas (OR 0.80/quintile increase, 95% CI 0.69-0.92) after being adjusted for age, typhoid fever vaccination, and home toilet type. Risk for exposure to Salmonella Typhi and its spatial distribution in Fiji are driven by environmental factors. Our findings can directly affect typhoid fever control efforts in Fiji.

Social mixing in Fiji: Who-eats-with-whom contact patterns and the implications of age and ethnic heterogeneity for disease dynamics in the Pacific Islands.

Empirical data on contact patterns can inform dynamic models of infectious disease transmission. Such information has not been widely reported from Pacific islands, nor strongly multi-ethnic settings, and few attempts have been made to quantify contact patterns relevant for the spread of gastrointestinal infections. As part of enteric fever investigations, we conducted a cross-sectional survey of the general public in Fiji, finding that within the 9,650 mealtime contacts reported by 1,814 participants, there was strong like-with-like mixing by age and ethnicity, with higher contact rates amongst iTaukei than non-iTaukei Fijians. Extra-domiciliary lunchtime contacts follow these mixing patterns, indicating the overall data do not simply reflect household structures. Inter-ethnic mixing was most common amongst school-age children. Serological responses indicative of recent Salmonella Typhi infection were found to be associated, after adjusting for age, with increased contact rates between meal-sharing iTaukei, with no association observed for other contact groups. Animal ownership and travel within the geographical division were common. These are novel data that identify ethnicity as an important social mixing variable, and use retrospective mealtime contacts as a socially acceptable metric of relevance to enteric, contact and respiratory diseases that can be collected in a single visit to participants. Application of these data to other island settings will enable communicable disease models to incorporate locally relevant mixing patterns in parameterisation.

Phenotypic and genotypic characteristics of ESBL and AmpC producing organisms associated with bacteraemia in Ho Chi Minh City, Vietnam.

BACKGROUND: Broad-spectrum antimicrobials are commonly used as empirical therapy for infections of presumed bacterial origin. Increasing resistance to these antimicrobial agents has prompted the need for alternative therapies and more effective surveillance. Better surveillance leads to more informed and improved delivery of therapeutic interventions, potentially leading to better treatment outcomes. METHODS: We screened 1017 Gram negative bacteria (excluding Pseudomonas spp. and Acinetobacter spp.) isolated between 2011 and 2013 from positive blood cultures for susceptibility against third generation cephalosporins, ESBL and/or AmpC production, and associated ESBL/AmpC genes, at the Hospital for Tropical Diseases in Ho Chi Minh City. RESULTS: Phenotypic screening found that 304/1017 (30%) organisms were resistance to third generation cephalosporins; 172/1017 (16.9%) of isolates exhibited ESBL activity, 6.2% (63/1017) had AmpC activity, and 0.5% (5/1017) had both ESBL and AmpC activity. E. coli and Aeromonas spp. were the most common organisms associated with ESBL and AmpC phenotypes, respectively. Nearly half of the AmpC producers harboured an ESBL gene. There was no significant difference (p > 0.05) between the antimicrobial resistance phenotypes of the organisms associated with community and hospital-acquired infections. CONCLUSION: AmpC and ESBL producing organisms were commonly associated with bloodstream infections in this setting, with antimicrobial resistant organisms being equally distributed between infections originating from the community and healthcare settings. Aeromonas spp., which was associated with bloodstream infections in cirrhotic/hepatitis patients, were the most abundant AmpC producing organism. We conclude that empirical monotherapy with third generation cephalosporins may not be optimum in this setting.

A cross-sectional seroepidemiological survey of typhoid fever in Fiji.

Fiji, an upper-middle income state in the Pacific Ocean, has experienced an increase in confirmed case notifications of enteric fever caused by Salmonella enterica serovar Typhi (S. Typhi). To characterize the epidemiology of typhoid exposure, we conducted a cross-sectional sero-epidemiological survey measuring IgG against the Vi antigen of S. Typhi to estimate the effect of age, ethnicity, and other variables on seroprevalence. Epidemiologically relevant cut-off titres were established using a mixed model analysis of data from recovering culture-confirmed typhoid cases. We enrolled and assayed plasma of 1787 participants for anti-Vi IgG; 1,531 of these were resident in mainland areas that had not been previously vaccinated against S. Typhi (seropositivity 32.3% (95%CI 28.2 to 36.3%)), 256 were resident on Taveuni island, which had been previously vaccinated (seropositivity 71.5% (95%CI 62.1 to 80.9%)). The seroprevalence on the Fijian mainland is one to two orders of magnitude higher than expected from confirmed case surveillance incidence, suggesting substantial subclinical or otherwise unreported typhoid. We found no significant differences in seropositivity prevalences by ethnicity, which is in contrast to disease surveillance data in which the indigenous iTaukei Fijian population are disproportionately affected. Using multivariable logistic regression, seropositivity was associated with increased age (odds ratio 1.3 (95% CI 1.2 to 1.4) per 10 years), the presence of a pit latrine (OR 1.6, 95%CI 1.1 to 2.3) as opposed to a septic tank or piped sewer, and residence in settlements rather than residential housing or villages (OR 1.6, 95% CI 1.0 to 2.7). Increasing seropositivity with age is suggestive of low-level endemic transmission in Fiji. Improved sanitation where pit latrines are used and addressing potential transmission routes in settlements may reduce exposure to S. Typhi. Widespread unreported infection suggests there may be a role for typhoid vaccination in Fiji, in addition to public health management of cases and outbreaks.

A universal genome sequencing method for rotavirus A from human fecal samples which identifies segment reassortment and multi-genotype mixed infection.

BACKGROUND: Genomic characterization of rotavirus (RoV) has not been adopted at large-scale due to the complexity of obtaining sequences for all 11 segments, particularly when feces are used as starting material. METHODS: To overcome these limitations, we developed a novel RoV capture and genome sequencing method combining commercial enzyme immunoassay plates and a set of routinely used reagents. RESULTS: Our approach had a 100% success rate, producing >90% genome coverage for diverse RoV present in fecal samples (Ct < 30). CONCLUSIONS: This method provides a novel, reproducible and comparatively simple approach for genomic RoV characterization and could be scaled-up for use in global RoV surveillance systems. TRIAL REGISTRATION (PROSPECTIVELY REGISTERED): Current Controlled Trials ISRCTN88101063 . Date of registration: 14/06/2012.

Evaluation of Luminex xTAG Gastrointestinal Pathogen Panel Assay for Detection of Multiple Diarrheal Pathogens in Fecal Samples in Vietnam.

Diarrheal disease is a complex syndrome that remains a leading cause of global childhood morbidity and mortality. The diagnosis of enteric pathogens in a timely and precise manner is important for making treatment decisions and informing public health policy, but accurate diagnosis is a major challenge in industrializing countries. Multiplex molecular diagnostic techniques may represent a significant improvement over classical approaches. We evaluated the Luminex xTAG gastrointestinal pathogen panel (GPP) assay for the detection of common enteric bacterial and viral pathogens in Vietnam. Microbiological culture and real-time PCR were used as gold standards. The tests were performed on 479 stool samples collected from people admitted to the hospital for diarrheal disease throughout Vietnam. Sensitivity and specificity were calculated for the xTAG GPP for the seven principal diarrheal etiologies. The sensitivity and specificity for the xTAG GPP were >88% for Shigellaspp.,Campylobacterspp., rotavirus, norovirus genotype 1/2 (GI/GII), and adenovirus compared to those of microbiological culture and/or real-time PCR. However, the specificity was low (∼60%) for Salmonella species. Additionally, a number of important pathogens that are not identified in routine hospital procedures in this setting, such as Cryptosporidiumspp. and Clostridium difficile, were detected with the GPP. The use of the Luminex xTAG GPP for the detection of enteric pathogens in settings, like Vietnam, would dramatically improve the diagnostic accuracy and capacity of hospital laboratories, allowing for timely and appropriate therapy decisions and a wider understanding of the epidemiology of pathogens associated with severe diarrheal disease in low-resource settings.

Clinical implications of reduced susceptibility to fluoroquinolones in paediatric Shigella sonnei and Shigella flexneri infections.

OBJECTIVES: We aimed to quantify the impact of fluoroquinolone resistance on the clinical outcome of paediatric shigellosis patients treated with fluoroquinolones in southern Vietnam. Such information is important to inform therapeutic management for infections caused by this increasingly drug-resistant pathogen, responsible for high morbidity and mortality in young children globally. METHODS: Clinical information and bacterial isolates were derived from a randomized controlled trial comparing gatifloxacin with ciprofloxacin for the treatment of paediatric shigellosis. Time-kill experiments were performed to evaluate the impact of MIC on the in vitro growth of Shigella and Cox regression modelling was used to compare clinical outcome between treatments and Shigella species. RESULTS: Shigella flexneri patients treated with gatifloxacin had significantly worse outcomes than those treated with ciprofloxacin. However, the MICs of fluoroquinolones were not significantly associated with poorer outcome. The presence of S83L and A87T mutations in the gyrA gene significantly increased MICs of fluoroquinolones. Finally, elevated MICs and the presence of the qnrS gene allowed Shigella to replicate efficiently in vitro in high concentrations of ciprofloxacin. CONCLUSIONS: We found that below the CLSI breakpoint, there was no association between MIC and clinical outcome in paediatric shigellosis infections. However, S. flexneri patients had worse clinical outcomes when treated with gatifloxacin in this study regardless of MIC. Additionally, Shigella harbouring the qnrS gene are able to replicate efficiently in high concentrations of ciprofloxacin and we hypothesize that such strains possess a competitive advantage against fluoroquinolone-susceptible strains due to enhanced shedding and transmission.

The Vietnam Initiative on Zoonotic Infections (VIZIONS): A Strategic Approach to Studying Emerging Zoonotic Infectious Diseases.

The effect of newly emerging or re-emerging infectious diseases of zoonotic origin in human populations can be potentially catastrophic, and large-scale investigations of such diseases are highly challenging. The monitoring of emergence events is subject to ascertainment bias, whether at the level of species discovery, emerging disease events, or disease outbreaks in human populations. Disease surveillance is generally performed post hoc, driven by a response to recent events and by the availability of detection and identification technologies. Additionally, the inventory of pathogens that exist in mammalian and other reservoirs is incomplete, and identifying those with the potential to cause disease in humans is rarely possible in advance. A major step in understanding the burden and diversity of zoonotic infections, the local behavioral and demographic risks of infection, and the risk of emergence of these pathogens in human populations is to establish surveillance networks in populations that maintain regular contact with diverse animal populations, and to simultaneously characterize pathogen diversity in human and animal populations. Vietnam has been an epicenter of disease emergence over the last decade, and practices at the human/animal interface may facilitate the likelihood of spillover of zoonotic pathogens into humans. To tackle the scientific issues surrounding the origins and emergence of zoonotic infections in Vietnam, we have established The Vietnam Initiative on Zoonotic Infections (VIZIONS). This countrywide project, in which several international institutions collaborate with Vietnamese organizations, is combining clinical data, epidemiology, high-throughput sequencing, and social sciences to address relevant one-health questions. Here, we describe the primary aims of the project, the infrastructure established to address our scientific questions, and the current status of the project. Our principal objective is to develop an integrated approach to the surveillance of pathogens circulating in both human and animal populations and assess how frequently they are exchanged. This infrastructure will facilitate systematic investigations of pathogen ecology and evolution, enhance understanding of viral cross-species transmission events, and identify relevant risk factors and drivers of zoonotic disease emergence.