Environmental Presence and Genetic Characteristics of Carbapenemase-Producing Enterobacteriaceae from Hospital Sewage and River Water in the Philippines.

This study aimed to evaluate the prevalence and genetic characteristics of carbapenemase-producing Enterobacteriaceae (CPE) in hospital sewage and river water in the Philippines, which has a typical tropical maritime climate. We collected 83 water samples from 7 hospital sewage and 10 river water sites. CPE were identified using CHROMagar mSuperCARBA, and Gram-negative strains were identified using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) or 16S rRNA gene sequencing. Resistance genes in Enterobacteriaceae strains were identified using PCR and DNA sequencing, and transferability of carbapenemase genes from the CPE was investigated with conjugation experiments. Genotyping was performed using multilocus sequence typing (MLST) for Escherichia coli and Klebsiella pneumoniae Out of 124 Enterobacteriaceae isolates, we identified 51 strains as CPE and divided these into 7 species, 11 E. coli, 14 Klebsiella spp., 15 Enterobacter spp., and 11 others, including 4 additional species. Conjugation experiments via broth mating and using E. coli J53 revealed that 24 isolates can transfer carbapenemase-encoding plasmids. MLST analysis showed that 6 of 11 E. coli isolates belonged to clonal complex 10 (CC10). Of 11 K. pneumoniae strains, 9 unique sequence types (STs) were identified, including ST147. Five types of carbapenemase genes were identified, with the most prevalent being NDM (n = 39), which is epidemic in clinical settings in the Philippines. E. coli CC10 and K. pneumoniae ST147, which are often detected in clinical settings, were the dominant strains. In summary, our results indicate that hospital sewage and river water are contaminated by CPE strains belonging to clinically important clonal groups.IMPORTANCE Carbapenemase-producing Enterobacteriaceae (CPE) cause severe health care-associated infections, and their increasing prevalence is a serious concern. Recently, natural ecosystems have been recognized as important reservoirs of antibiotic resistance genes. We investigated the prevalence and genetic characteristics of CPE isolated from the environment (hospital sewage and river water) in the Philippines and found several CPE, including Escherichia coli and other species, with different carbapenemases. The most prevalent carbapenemase gene type was NDM, which is endemic in clinical settings. This study revealed that isolates belonging to carbapenemase-producing E. coli CC10 and K. pneumoniae sequence type 147 (ST147), which are often detected in clinical settings, were dominant in the natural environment. Our work here provides a report on the presence and characteristics of CPE in the environment in the Philippines and demonstrates that both hospital sewage and river water are contaminated by CPE strains belonging to clinically important clonal groups.

Molecular characterization of Vibrio cholerae O1 isolates obtained from outbreaks in the Philippines, 2015-2016.

Introduction. The Philippines, comprising three island groups, namely, Luzon, Visayas and Mindanao, experienced an increase in cholera outbreaks in 2016. Previous studies have shown that Vibrio cholerae isolates obtained from the Philippines are novel hybrid El Tor strains that have evolved in the country and are clearly distinct from those found in Mozambique and Cameroon.Gap statement. The characterization of the strains isolated from outbreaks has been limited to phenotypic characteristics, such as biochemical and serological characteristics, in most previous studies.Aim. We performed multilocus variable-number tandem repeat (VNTR) analysis (MLVA) for V. cholerae isolates obtained from 2015 to 2016 to further characterize and understand the emergence and dissemination of the strains in the Philippines.Methodology. A total of 139 V. cholerae O1 Ogawa biotype El Tor isolates were obtained from the Philippines during diarrhoeal outbreaks in 18 provinces between 2015 and 2016. VNTR data were analysed to classify the MLVA profiles where the large-chromosome types (LCTs) were applied for grouping.Results. We identified 50 MLVA types among 139 isolates originating from 18 provinces, and 14 LCTs. The distribution of the LCTs was variable, and a few were located in specific areas or even in specific provinces. Based on eBURST analysis, 99 isolates with 7 LCTs and 32 MLVA types belonged to 1 group, suggesting that they were related to each other. LCT A was predominant (n=67) and was isolated from Luzon and Visayas. LCT A had 14 MLVA types; however, it mostly emerged during a single quarter of a year. Eight clusters were identified, each of which involved specific MLVA type(s). The largest cluster involved 23 isolates showing 3 MLVA types, 21 of which were MLVA type A-14 isolated from Negros Occidental during quarter 4 of 2016. Comparative analysis showed that almost all isolates from the Philippines were distinct from those in other countries.Conclusions. The genotypic relationship of the V. cholerae isolates obtained during outbreaks in the Philippines was studied, and their emergence and dissemination were elucidated. MLVA revealed the short-term dynamics of V. cholerae genotypes in the Philippines.

HLA-A*33:01 as protective allele for severe dengue in a population of Filipino children.

Dengue virus infection is a leading cause of morbidity among children in the Philippines in recent years. In order to investigate the association of HLA Class I and II alleles and dengue disease severity in a cohort of Filipino children, we performed a case control study in 2 hospitals in Metro Manila from June 2008 to December 2009. A total of 250 laboratory confirmed dengue patients and 300 healthy individuals aged 5 to 15 years old were typed for HLA-A, B and DRB1 alleles. The frequency of HLA-A*33:01 was significantly decreased in severe dengue (DHF/ DSS; Pc = 0.0016)) and DSS (Pc = 0.0032) compared to the background population. These findings support a previous study that this allele may confer protection against the severe form of dengue and provide the first evidence of HLA association with dengue in the Philippines. Future studies should be directed in investigating the possible mechanisms of protection.

Alpha tryptase allele of Tryptase 1 (TPSAB1) gene associated with Dengue Hemorrhagic Fever (DHF) and Dengue Shock Syndrome (DSS) in Vietnam and Philippines.

We previously reported, significantly higher levels of Chymase and Tryptase in early stage plasma of DSS patients prior to the occurrence of shock suggesting a possible role of mast cells in dengue pathogenesis. To further investigate, we analyzed CMA1 promoter SNP (rs1800875) and TPSAB1 gene alleles, which encode the Human Chymase and α- and ß- tryptase 1 enzymes respectively, for susceptibility to Dengue Hemorrhagic Fever (DHF) and Dengue Shock Syndrome (DSS) in patients from hospitals in Vietnam (Ho Chi Minh City and Vinh Long) and the Philippines. While the CMA1 promoter SNP (rs1800875) was not associated with DHF/DSS, the homozygous form of α-tryptase allele was associated with DSS patients in Vinh Long and the Philippines (OR=3.52, p<0.0001; OR=3.37, p<0.0001, respectively) and with DHF in Ho Chi Minh City (OR=2.54, p=0.0084). Also, a statistically significant association was observed when DHF and DSS were combined in Vinh Long (OR=1.5, p=0.034) and the Philippines (OR=2.36, p=0.0004); in Ho Chi Minh City when DHF and DSS were combine an association was observed, but it was not statistically significant (OR=1.5, p=0.0505). Therefore, the α-tryptase might have a possible effect on the susceptibility to severe form of Dengue infection.