ナノ流路技術を用いたリアルタイムPCRシステムによる肺炎球菌血清型分子診断法の開発とその応用について

長崎大学学位論文 [学位記番号]博（医歯薬）甲第730号 [学位授与年月日]平成26年12月3

Novel hemagglutinating, hemolytic and cytotoxic activities of the intermediate subunit of Entamoeba histolytica lectin

Galactose and N-acetyl-D-galactosamine (Gal/GalNAc) inhibitable lectin of Entamoeba histolytica, a common protozoan parasite, has roles in pathogenicity and induction of protective immunity in mouse models of amoebiasis. The lectin consists of heavy (Hgl), light (Lgl), and intermediate (Igl) subunits. Hgl has lectin activity and Lgl does not, but little is known about the activity of Igl. In this study, we assessed various regions of Igl for hemagglutinating activity using recombinant proteins expressed in Escherichia coli. We identified a weak hemagglutinating activity of the protein. Furthermore, we found novel hemolytic and cytotoxic activities of the lectin, which resided in the carboxy-terminal region of the protein. Antibodies against Igl inhibited the hemolytic activity of Entamoeba histolytica trophozoites. This is the first report showing hemagglutinating, hemolytic and cytotoxic activities of an amoebic molecule, Igl

A major dengue epidemic in 2022 in Nepal: need of an efficient early-warning system

Dengue has become a recurrent and growing threat to public health in Nepal. The epidemic in 2022 was the largest ever reported, with cases being reported in all of the country’s seven provinces and 77 districts. Despite the establishment of an early-warning and reporting system (EWARS) in 1997, the lack of clear criteria for alarm signals and outbreak definitions in national guidelines delayed the epidemic declaration in 2022, which resulted in an increased number of cases and fatalities. For this article, we analyzed national data from previous years, which demonstrate that an epidemic could have been declared early in July, and that that would have resulted in fewer cases and fatalities if clear criteria for outbreak declarations had also been put in place. We also reviewed the existing national guidelines for dengue prevention and control, and propose recommendations to improve their implementation, particularly with regard to vector control measures. This article also highlights the need for a coordinated effort between multisector stakeholders, strengthened disease surveillance systems, and the establishment of predefined alarm signals and epidemic declaration criteria so that future epidemics are identified in a timely manner. The early outbreak warning system can potentially prevent future large outbreaks and minimize their negative impacts on the country’s health systems and economy

The overlap of accessory virulence factors and multidrug resistance among clinical and surveillance Klebsiella pneumoniae isolates from a neonatal intensive care unit in Nepal: a single-centre experience in a resource-limited setting

Background: There is a lack of data on the characteristics of overlap between acquired antimicrobial resistance and virulence factors in Klebsiella pneumoniae in high-risk settings, especially with the inclusion of surveillance isolates along with the clinical. We investigated K. pneumoniae isolates, from a neonatal intensive care unit (NICU) in Nepal, for the presence of both accessory virulence factors and acquired antimicrobial resistance. Methods: Thirty-eight clinical and nineteen surveillance K. pneumoniae isolates obtained between January 2017 and August 2022 in the NICU of Siddhi Memorial Hospital, Bhaktapur, Nepal were investigated with antimicrobial susceptibility testing, PCR-based detection of β-lactamases and virulence factors, and genetic similarity by ERIC–PCR. Results: K. pneumoniae was found positive in 37/85 (43.5%) blood culture-positive neonatal bloodstream infections, 34/954 (3.6%) patient surveillance cultures, and 15/451 (3.3%) environmental surveillance samples. Among 57 isolates analyzed in this study, we detected multidrug resistance in 37/57 (64.9%), which was combined with at least one accessory virulence factor in 21/37 (56.8%). This overlap was mostly among β-lactamase producing isolates with accessory mechanisms of iron acquisition. These isolates displayed heterogenous ERIC–PCR patterns suggesting genetic diversity. Conclusions: The clinical significance of this overlap between acquired antimicrobial resistance and accessory virulence genes in K. pneumoniae needs further investigation. Better resource allocation is necessary to strengthen infection prevention and control interventions in resource-limited settings

The causes of bacterial bloodstream infections and antimicrobial resistance patterns in children attending a secondary care hospital in Bhaktapur, Nepal, 2017–2022: a retrospective study

Objectives: Data on antimicrobial resistance (AMR) among children in Nepal are limited. Here we have characterized the causes of bacterial bloodstream infections (BSIs), antimicrobial resistance patterns and the mechanisms of β-lactamase production in Enterobacterales among children attending outpatient and inpatient departments of a secondary care paediatric hospital in Nepal. Methods: We retrospectively collected demographic and clinical data of culture-proven bacterial BSIs between January 2017 and December 2022 among children <18 years attending a 50-bedded paediatric hospital. Stored isolates were subcultured for antimicrobial susceptibility testing against commonly used antimicrobials. Enterobacterales displaying non-susceptibility to β-lactams were phenotypically and genotypically investigated for ESBLs, plasmid-mediated AmpC (pAmpC) β-lactamases and carbapenemases. Results: A total of 377 significant bacteria were isolated from 27 366 blood cultures. Among 91 neonates with a BSI, Klebsiella pneumoniae (n = 39, 42.4%), Pseudomonas aeruginosa (n = 15, 16.3%) and Acinetobacter baumannii complex (n = 13, 14.1%) were most common. In the non-neonates, 275/285 (96.5%) infections were community-acquired including Staphylococcus aureus (n = 89, 32.4%), Salmonella Typhi (n = 54, 19.6%) and Streptococcus pneumoniae (n = 32, 11.6%). Among the 98 S. aureus, 29 (29.6%) were methicillin-resistant Staphylococcus aureus. K. pneumoniae and Escherichia coli demonstrated non-susceptibility to extended-spectrum cephalosporins and carbapenems in both community and hospital-acquired cases. For E. coli and K. pneumoniae, blaCTX-M (45/46), blaEBC (7/10) and blaOXA-48 (5/6) were common among their respective groups. Conclusions: We determined significant levels of AMR among children attending a secondary care paediatric hospital with BSI in Nepal. Nationwide surveillance and implementation of antimicrobial stewardship policies are needed to combat the challenge imposed by AMR

Risk factors for wasting among hospitalised children in Nepal

Background: Malnutrition has various adverse effects in children. This study aimed to determine risk factors for malnutrition among hospitalised children, changes in nutritional status at admission and discharge and effects of use of systematic anthropometric measurement in identification of malnutrition.Methods: We enrolled 426 children, aged between 6 months and 15 years, admitted to Siddhi Memorial Hospital, Bhaktapur, Nepal, from November 2016 to June 2017. Anthropometric measurements were performed at the time of admission and discharge. Risk factors were assessed by multivariable logistic regression models.Results: Median age of children was 26 months (IQR: 13–49), and males were 58.7%. The prevalence of wasting was 9.2% (39/426) at admission and 8.5% (36/426) at discharge. Risk factors associated with wasting at admission were ethnic minority (aOR: 3.6, 95% CI 1.2–10.8), diarrhoeal diseases (aOR = 4.0; 95% CI 1.3–11.8), respiratory diseases (aOR: 3.4, 95% CI 1.4–8.1) and earthquake damage to house (aOR = 2.6; 95% CI 1.1–6.3). Clinical observation by care providers identified only 2 out of 112 malnutrition cases at admission and 4 out of 119 cases at discharge that were detected by the systematic anthropometric measurement.Conclusions: Ethnic minority, diarrhoeal diseases, respiratory infections and house damage due to the earthquake were risk factors associated with wasting. Systematic anthropometric examination can identify significantly more malnourished children than simple observation of care providers

Prevalence of vitamin D deficiency in pregnant women and their babies in Bhaktapur, Nepal.

BACKGROUND: Vitamin D deficiency has been observed worldwide in pregnant women and their newborns. Maternal vitamin D deficiency can lead to deficiency in their newborn baby and has been linked with various complications during pregnancy and delivery. There is risk of premature delivery and it is associated with high neonatal mortality. METHODS: Seventy-nine pregnant women who were admitted to the Siddhi Memorial Hospital for delivery and their newborn babies were enrolled in the study. Maternal blood samples were taken before delivery while umbilical cord blood samples of their babies were taken after delivery. Serum vitamin D level and calcium level were assessed by fluorescence immunoassay using Ichromax vitamin D kit and endpoint method, respectively in the Siddhi Memorial Hospital laboratory. RESULTS: Mean +/- SD serum vitamin D and calcium levels in pregnant mother before delivery were 14.6 +/- 8.5 ng/ml and 8.0 +/- 0.5 mg/dl, respectively, and in the cord blood were 25.7 +/- 11.2 ng/ml and 8.6 +/- 0.9 mg/dl, respectively. Eighty-one percent of the mothers and 35.8% of their babies were found to have vitamin D deficiency. Although 97.5% of the pregnant women were taking calcium supplementation, serum calcium was found lower than the normal reference value in 67% of the pregnant women and 64.2% of their babies. There were a linear relationship between the maternal and baby's serum vitamin D (P < 0.001) and calcium (P < 0.001) levels. CONCLUSION: There is high prevalence of vitamin D and calcium deficiency in pregnant mothers and newborn babies in Bhaktapur, Nepal. Pregnant women need to be supplemented with adequate amounts of these nutrients

A description of a pre-emptive typhoid Vi capsular polysaccharide vaccination campaign after the 2015 earthquake in Nepal and vaccine effectiveness evaluation

Background: A 7.8 R scale earthquake hit Nepal in April 2015 and caused about 9000 deaths along with damage to infrastructure, including the water and sewage system. Bhaktapur was one of the highly affected districts. A typhoid vaccination campaign (pre-emptive) was carried out among children who were living in the temporary shelters in this district. The assessment of vaccine effectiveness after a pre-emptive typhoid vaccine campaign following an earthquake has previously not been attempted in Nepal. Objective: To describe the pre-emptive typhoid Vi capsular polysaccharide vaccination campaign and an evaluation of the vaccine effectiveness. Methods: We conducted a pre-emptive typhoid Vi capsular polysaccharide vaccination campaign among children between 2 and 15 years of age dwelling in 23 temporary shelters in Bhaktapur district after the earthquake. Surveillance of clinical typhoid was carried out from 2014 to 2017 in Siddhi Memorial Hospital, the only hospital for children in the district. We calculated vaccine effectiveness using a case–control study design (clinical typhoid as cases and chest x-ray confirmed pneumonia as controls). Results: Three thousand nine hundred sixteen children of age 2–15 years residing in the 23 temporary shelters in Bhaktapur received the typhoid Vi capsular polysaccharide vaccine between July and December 2015. 2193 children of age 2–15 years were admitted to the hospital during the study period and 260 (11.9%) were diagnosed with clinical typhoid. The numbers of children admitted with clinical typhoid decreased over the study period (105 in 2014 and 47 in 2017; P = 0.001). Overall vaccine effectiveness was calculated at 52% (95% CI −46 to 85%), and it was 87% (95% CI −25 to 99) among children less than 5 years of age. Conclusions: We successfully conducted a pre-emptive vaccination campaign against typhoid after the 2015 Nepal earthquake. The pre-emptive vaccination campaign appeared to be more effective among children less than 5 years of age. Further studies are needed to assess the effectiveness of pre-emptive use of typhoid vaccines in the emergency situations. We highlight the challenges of calculating vaccine effectiveness of a typhoid vaccine in an emergency setting

Ambient PM2.5 and Daily Hospital Admissions for Acute Respiratory Infections: Effect Modification by Weight Status of Child

The high level of ambient particulate matter in many developing countries constitutes a major health burden, but evidence on its impact on children’s health is still limited in these regions. We conducted a time-stratified case-crossover analysis to quantify the short-term association between fine particulate matter (PM2.5) and hospital admissions due to acute respiratory infections (ARI) among children in Bhaktapur district, Nepal, and to investigate the potential modification of the effect by nutritional characteristic. We analyzed 258 children admitted to the pediatric hospital for ARI between February 2014 to February 2015. We observed evidence of increased risk on the same (lag 0) and preceding day (lag 1). The cumulative estimate of their average (lag 01) suggested each 10 μg/m3 increase in PM2.5 was associated with a relative risk (RR) of 1.16 (95% confidence interval [CI]: 1.02–1.31). The strongest evidence from a stratified analysis of three categories of weights was observed in the overweight group (RR: 1.77; 95% CI: 1.17–2.69) at lag 01, while the estimates for the normal weight and underweight groups were closer to the non-stratified estimates for all-ARI cases. The findings suggests that pediatric ARI is an important morbidity associated with inhalable PM2.5 and that more research is needed to elucidate and validate the observed dissimilarity by weight