Isolation and characterization of influenza C viruses in the Philippines and Japan.

From November 2009 to December 2013 in the Philippines, 15 influenza C viruses were isolated, using MDCK cells, from specimens obtained from children with severe pneumonia and influenza-like illness (ILI). This is the first report of influenza C virus isolation in the Philippines. In addition, from January 2008 to December 2013, 7 influenza C viruses were isolated from specimens that were obtained from children with acute respiratory illness (ARI) in Sendai city, Japan. Antigenic analysis with monoclonal antibodies to the hemagglutinin-esterase (HE) glycoprotein showed that 19 strains (12 from the Philippines and 7 from Japan) were similar to the influenza C virus reference strain C/Sao Paulo/378/82 (SP82). Phylogenetic analysis of the HE gene showed that the strains from the Philippines and Japan formed distinct clusters within an SP82-related lineage. The clusters that included the Philippine and Japanese strains were shown to have diverged from a common ancestor around 1993. In addition, phylogenetic analysis of the internal genes showed that all strains isolated in the Philippines and Japan had emerged through reassortment events. The composition of the internal genes of the Philippine strains was different from that of the Japanese strains, although all strains were classified into an SP82-related lineage by HE gene sequence analysis. These observations suggest that the influenza C viruses analyzed here had emerged through different reassortment events; however, the time and place at which the reassortment events occurred were not determined.

Pneumonia among young infants in rural Southeast Asia (Bohol Island, Philippines).

OBJECTIVE: To develop a clinical algorithm that can be used to identify pneumonia deaths in young infants in developing countries and estimate the disease burden in this population. PATIENTS AND METHODS: Infants younger than 60 days hospitalized with signs of severe pneumonia who underwent clinical, microbiologic and radiological evaluation were the subjects. Stepwise logistic regression and subtractive iterative process were used to derive the algorithm. RESULTS: Three-hundred and one infants had either clinical or radiographic pneumonia. The case fatality rate for 185 infants with radiographic pneumonia was 21%vs. 5% for clinical pneumonia. Age below 7 days was associated with an increased risk of dying. Among 7- to 59-day-old infants, poor feeding, cyanosis and absence of crackles were predictors of death from pneumonia. Using logistic regression, an algorithm consisting of any one of three clinical signs (cyanosis, poor feeding and abnormally sleepy) was developed in infants aged 7-59 days; 80% of deaths and 50% of those with radiographic pneumonia have at least one of these signs. It performed better than both the WHO case management algorithm and the IMCI algorithm. CONCLUSION: Radiographic pneumonia is a common and serious infection among infants below 2 months old in the Philippines. Cyanosis, poor feeding and abnormal sleepiness are simple signs that can be used by health workers to identify seriously ill infants who are most likely to die from pneumonia.

Molecular epidemiology of Bordetella pertussis in the Philippines in 2012-2014.

OBJECTIVES: The present study was designed to determine the genotypes of circulating Bordetella pertussis in the Philippines by direct molecular typing of clinical specimens. METHODS: Nasopharyngeal swabs (NPSs) were collected from 50 children hospitalized with pertussis in three hospitals during 2012-2014. Multilocus variable-number tandem repeat analysis (MLVA) was performed on the DNA extracts from NPSs. B. pertussis virulence-associated allelic genes (ptxA, prn, and fim3) and the pertussis toxin promoter, ptxP, were also investigated by DNA sequence-based typing. RESULTS: Twenty-six DNA extracts yielded a complete MLVA profile, which were sorted into 10 MLVA types. MLVA type 34 (MT34), which is rare in Australia, Europe, Japan, and the USA, was the predominant strain (50%). Seven MTs (MT29, MT32, MT33, and MT283-286, total 42%) were single-locus variants of MT34, while two (MT141 and MT287, total 8%) were double-locus variants of MT34. All MTs had the combination of virulence-associated allelic genes, ptxP1-ptxA1-prn1-fim3A. CONCLUSIONS: The B. pertussis population in the Philippines comprises genetically related strains. These strains are markedly different from those found in patients from other countries where acellular pertussis vaccines are used. The differences in vaccine types between these other countries and the Philippines, where the whole-cell vaccine is still used, may select for distinct populations of B. pertussis.

Molecular characteristics of extended-spectrum ß-lactamases in clinical isolates of Enterobacteriaceae from the Philippines.

ß-Lactamases, including extended-spectrum ß-lactamases (ESBLs) and AmpC ß-lactamases, are major resistance mechanisms of Enterobacteriaceae. Emergence of plasmid-mediated quinolone resistance (PMQR) determinants in ESBL-producing isolates poses a global threat. The molecular characterisitcs of ESBL and PMQR determinants in the Philippines are not well characterized. In this study, we investigated ESBLs and AmpC ß-lactamases in clinical isolates of Enterobacteriaceae from the Philippines, and analyzed the association between ESBL and PMQR genes. A total of 91 amoxicilin non-susceptible Enterobacteriaceae were collected at the Research Institute for Tropical Medicine of the Philippines from 2006 to 2008. AmpC- or ESBL-producing isolates were screened by detecting a zone diameter for cefoxitin ≤ 14 mm or cefpodoxime ≤ 20 mm, respectively. Possible ESBL-producing strains were assessed by the ESBL confirmation test of the Clinical and Laboratory Standards Institute. PCR and sequencing were performed to detect the ESBL and PMQR genes. The number of ESBL-producers and AmpC-producers confirmed phenotypically was 17 (18.7%) and 61 (67.0%), respectively. Of 17 phenotypic ESBL-producers, 14 isolates had ESBL genes, including 6 of Escherichia coli, 3 of Enterobacter cloacae, 2 of Enterobacter aerogenes, 2 of Klebsiella pneumoniae, and 1 of Klebsiella ozaenae. Among these isolates, there were 13, 4, and 12 with bla(CTX-M), bla(SHV), and bla(TEM), respectively. Of the bla(CTX-M)-positive isolates, bla(CTX-M-15) shows the highest prevalence, followed by bla(CTX-M-3) and bla(CTX-M-14). Of 14 ESBL-producers identified by PCR, 4, 6, and 7 isolates were positive for qnrB, qnrS, and aac(6')-Ib-cr, respectively. The frequency of aac(6')-Ib-cr positivity was significantly higher among CTX-M-15-producing isolates. Thus, we identified bla(CTX-M), aac(6')-Ib-cr, and qnr in ESBL-producing Enterobacteriaceae from the Philippines, and revealed a significant association between bla(CTX-M-15) and aac(6')-Ib-cr. Local epidemiological data are important for implementing appropriate antimicrobial therapy and effective infection control measures. Continuous monitoring of antimicrobial resistance genes in the Philippines will be required.

Central nervous system infection is an important cause of death in underfives hospitalised with World Health Organization (WHO) defined severe and very severe pneumonia.

Over 6 years, 1667 children aged 2-59 months admitted for pneumonia [1287 severe and 380 very severe] were studied. The case fatality rate (CFR) in children with severe pneumonia was 2.1% and 14.3% with CNS infection, with very severe pneumonia the CFR was 18.9%, 10.4% in those with hypoxemia and 43.6% with CNS infection. High CFRs were associated with CNS infection and inability to drink/cyanosis. The appropriate management of children with very severe pneumonia should include cerebrospinal fluid examination, oxygen monitoring and possibly ventilated support, suggesting that these are minimal standards of care at the district hospital.

Serious community-acquired paediatric infections in rural Asia (Bohol Island, Philippines): bacterial meningitis in children less than 5 years of age.

This paper describes the clinical profile and aetiology of bacterial meningitis in infants and children less than 5 y old admitted to a rural general hospital in the Philippines. A total of 989 infants and children 0-59 months old with suspected meningitis using a standardized guideline based on clinical signs and symptoms were prospectively enrolled from April 1994 to May 2000. Blood and CSF were drawn on admission for culture, antigen testing and cell count. All had blood cultures and 623 (63%) had CSF samples. Bacterial aetiology was found in 54 (5%). The most common bacterial pathogens were H. influenzae type b (Hib) (20, 37%) and S. pneumoniae (Pnc) (10, 18%). All of the Hib infections and 8 (80%) Pnc infections were in infants less than 1 y old. 12 (22%) of the subjects with bacterial meningitis died. All strains of S. pneumoniae and H. influenzae were sensitive to chloramphenicol, cotrimoxazole and ampicillin. In conclusion, S. pneumoniae and H. influenzae type b are the most common aetiological agents of bacterial meningitis in a rural area in the Philippines, and occur especially in infants less than 1 y old. Aetiological agents were susceptible to the currently recommended antimicrobial agents.