Viral burden and diversity in acute respiratory tract infections in hospitalized children in wet and dry zones of Sri Lanka.

The present study was done to identify the viral diversity, seasonality and burden associated with childhood acute respiratory tract infection (ARTI) in Sri Lanka. Nasopharyngeal aspirates (NPA) of hospitalized children (1 month-5 years) with ARTI were collected in 2 centers (wet and dry zones) from March 2013 to August 2014. Respiratory viral antigen detection by immunofluorescence assay (IFA) was used to identify the infecting viruses. IFA negative 100 NPA samples were tested for human metapeumovirus (hMPV), human bocavirus and corona viruses by polymerase chain reaction. Of the 443 and 418 NPAs, 37.2% and 39.4% were positive for any of the 8 different respiratory viruses tested from two centers studied. Viral co-infection was detected with respiratory syncytial virus (RSV) in both centers. Peak viral detection was noted in the wet zone from May-July 2013 and 2014 and in the dry zone from December-January 2014 suggesting a local seasonality for viral ARTI. RSV showed a clear seasonality with a direct correlation of monthly RSV infections with rainy days in the wet zone and an inverse correlation with temperature in both centers. The case fatality rate was 2.7% for RSV associated ARTI. The overall disability adjusted life years was 335.9 and for RSV associated ARTI it was 241.8. RSV was the commonly detected respiratory virus with an annual seasonality and distribution in rainy seasons in the dry and wet zones of Sri Lanka. Identifying the virus and seasonality will contribute to employ preventive measures and reduce the empirical use of antibiotics in resource limited settings.

Dengue virus co-infections with multiple serotypes do not result in a different clinical outcome compared to mono-infections.

Circulation of multiple dengue virus (DENV) serotypes in a locale has resulted in individuals becoming infected with mixed serotypes. This research was undertaken to study the clinical presentation, presence of DENV serotypes and serological characteristics of DENV infected patients with co-infections from three Provinces of Sri Lanka where DENV-1 and -2 predominated during the study. A reverse transcription polymerase chain reaction was performed on 1249 patient samples and 301 were positive for DENV (24.1%). DENV-1 was the predominant serotype detected in 137 (45.51%) followed by DENV-2 in 65 (21.59%), DENV-3 in 59 (19.6%) and DENV-4 in 4 (1.32%) patients with mono-infections. Thirty-three patients (10.96%) had DENV co-infections with two or more serotypes. The highest number of co-infections was noted between DENV-1 and DENV-2 (57.57%) suggesting co-infection is driven by the frequency of the circulating serotypes. Platelet counts were significantly higher in DENV co-infected patients although clinical disease severity or white blood cell count, packed cell volume or viraemia were not significantly different in the co-infected compared to the mono-infected patients. Thus co-infection with multiple DENV serotypes does occur but with the exception of improved platelet counts in co-infected patients, there is no evidence that clinical or laboratory measures of disease are altered.

Salinity tolerant Aedes aegypti and Ae. albopictus-Infection with dengue virus and contribution to dengue transmission in a coastal peninsula.

BACKGROUND & OBJECTIVES: Aedes aegypti and Ae. albopictus are major arboviral vectors that are considered to lay eggs, and undergo preimaginal development only in fresh water collections. However, recently they have been also shown to develop in coastal brackish water habitats. The ability of the biologically variant salinity-tolerant Aedes vectors to transmit arboviral diseases is unknown. We therefore, investigated the infection of salinity-tolerant Aedes mosquitoes with dengue virus (DENV) and analysed dengue incidence and rainfall data to assess the contribution of salinity-tolerant Aedes vectors to dengue transmission in the coastal Jaffna peninsula in Sri Lanka. METHODS: Brackish and fresh water developing female Ae. aegypti and Ae. albopictus were tested for their ability to become infected with DENV through in vitro blood feeding and then transmit DENV vertically to their progeny. An immunochromatographic test for the NS1 antigen was used to detect DENV. Temporal variation in dengue incidence in relation to rainfall was analysed for the peninsula and other parts of Sri Lanka. RESULTS: Aedes aegypti and Ae. albopictus developing in brackish water, became infected with DENV through in vitro blood feeding and the infected mosquitoes were able to vertically transmit DENV to their progeny. Monsoonal rainfall was the discernible factor responsible for the seasonal increase in dengue incidence in the peninsula and elsewhere in Sri Lanka. INTERPRETATION & CONCLUSION: Fresh water Aedes vectors are main contributors to the increased dengue incidence that typically follows monsoons in the Jaffna peninsula and elsewhere in Sri Lanka. It is possible however, that brackish water-developing Aedes constitute a perennial reservoir for DENV to maintain a basal level of dengue transmission in coastal areas of the peninsula during the dry season, and this supports increased transmission when monsoonal rains expand populations of fresh water Aedes.

Coronaviruses in guano from Pteropus medius bats in Peradeniya, Sri Lanka.

Bats are a unique group of mammals well suited to be hosts for emerging viruses. With current rates of deforestation and urbanization, redistribution of bat habitats to urban and suburban areas may bring bats into closer contact with livestock and humans. Common flying fox, Pteropus medius (previously known as Pteropus giganteus), forms large communal roosts on treetops, often in close proximity to human habitation in Sri Lanka. This report describes the detection of coronavirus RNA in P. medius bat guano collected in Peradeniya, Sri Lanka. These viruses had >97% nucleotide identity with coronaviruses detected in Cynopterus sphinx, Scotophilus heathii and S. kuhlii bats in Thailand. Pteropus medius is widespread in Asia and appears to excrete group D coronaviruses, which are hitherto confined to bats; however, these findings may have public health implications in the future.

A case series on common cold to severe bronchiolitis and pneumonia in children following human metapneumovirus infection in Sri Lanka.

OBJECTIVES: The prevalence of hMPV infections in Sri Lanka has not been reported and here we report a case series of hMPV infection in children less than 5 years. Patients with ARTI were included from Teaching Hospital, Anuradhapura from March 2013 to August 2014. Indirect fluorescence assay was performed on nasopharyngeal aspirates for the identification of respiratory viruses [respiratory syncytial virus (RSV), parainfluenza virus 1, 2 and 3, influenza A and B and hMPV]. Moreover, reverse transcriptase-polymerase chain reaction was done to further confirm the hMPV infection. RESULTS: In this case series, hMPV infection showed a range of respiratory symptoms from common cold to life threatening lower respiratory tract infections with varying severity. In some cases, the clinical presentation of hMPV infection was similar to the ARTI caused by RSV. hMPV co-infections with of RSV have also been seen in some cases of ARTI. A child delivered through cesarean section and birth order > 3 has an Odds ratio of 3.5 and 4.3 (95% CI) for developing co-infection with RSV compared to hMPV mono-infections. Lack of diagnostic facilities to identify the viral aetiology has contributed to the use of antibiotics indicating the need for establishing viral diagnostic facilities in the country.

History and current trends in influenza virus infections with special reference to Sri Lanka.

The World Health Organization (WHO) estimates that approximately one billion people are infected and up to 500,000 people die from influenza each year in the world. Influenza is considered to be the greatest killer of the human populations, due to the 1918 Spanish flu, which killed millions around the world. Despite the effective treatment available against influenza, it still contributes to significant morbidity and mortality. Currently circulating influenza strains in humans include influenza A (H1N1)pdm09, influenza A (H3N2) and influenza B viruses, (B/Victoria and B/Yamagata). Influenza has been prevalent in Sri Lanka from 1969, since then it continued to cause morbidity and mortality in children and adults. The current global influenza surveillance network monitors the global influenza activity through WHO collaborating centres. The Medical Research Institute monitors and diagnoses influenza cases in the country as part of the WHO network laboratories. Vaccinations to high risk groups and antiviral therapy for the successful prevention of influenza have been practiced in Sri Lanka. This review highlights the impact of influenza on public health in Sri Lanka including the historical aspects, current diagnostic practices and prevention approaches in high risk individuals in the country.

Genotypes of hepatitis B virus identified in patients tested prior to endoscopy from a Teaching Hospital in the Central Province of Sri Lanka.

The present study was carried out to identify the hepatitis B virus (HBV) genotypes in six patients attending the surgical clinic who were positive for hepatitis B surface antigen. DNA was extracted from the serum of patients and subjected to a modified nested PCR to detect a 585 bp region within the S gene of the HBV genome. Positive PCR products were purified and sequenced via a cycle sequencing method. The sequence data were analyzed with reference sequences in the HepSEQ database to identify the particular HBV genotype of the samples. Nested PCR for the S gene of the HBV genome was positive in 2 out of 6 samples. The genotyping and sequence analysis of the PCR products showed HBV genotype A with a homology of 98% to the reference sequences in the HepSeq database.

A low cost rapid urease test to detect Helicobacter pylori infection in resource limited settings.

The aim of the study was to evaluate the suitability of a modified one minute rapid urease test (one day rapid urease test) as a low cost H. pylori detection method. A sample of 205 patients clinically suspected of having H. pylori infection was tested. One day rapid urease test and histology based H. pylori tests (the gold standard) were performed on endoscopic antral biopsies. There were 6 true positives, 191 true negatives, 8 false positives and zero false negatives. The sensitivity, specificity and positive (PPV) and negative predictive values (NPV) of the test were 100%, 96%, 42.9%, and 100% respectively. The cost per patient was 0.3US$. High sensitivity, specificity and NPV, low cost and simplicity of method were the advantages of the test and the main limitation was low PPV. Hence, one day rapid urease test can be considered as a suitable low cost method to detect H. pylori infection in resource limited settings.

Prevalence of Helicobacter pylori in benign gastric ulcers in a cohort of Sri Lankan patients.

Helicobacter pylori prevalence is decreasing globally and prevalence of non H. pylori gastric ulcers is increasing. The following study was conducted to assess the prevalence of H. pylori in benign gastric ulcers in a sample of Sri Lankan patients. This was a cross-sectional study of 59 dyspeptic patients with benign gastric ulcers. Multiple endoscopic gastric biopsies were obtained and histology, immunohistochemistry and polymerase chain reaction were performed for H. pylori detection. An immunochromatography assay was performed to detect blood anti H. pylori antibodies. Four (6.8%) were positive for H. pylori. Therefore, it is likely that most benign gastric ulcers are of non-H. pylori aetiology.

Prevalence and identification of Cryptosporidium species in paediatric patients with diarrhoea.

OBJECTIVES: To determine the prevalence of Cryptosporidium infection in children with diarrhoea, identify associated factors and identify the parasite using Polymerase Chain Reaction (PCR). METHODS: A total of 138 diarrhoeic faecal samples were collected between August 2011 and February 2013, from children under 12 years of age, admitted to paediatric units of Teaching Hospitals, Kandy and Peradeniya, Sirimawo Bandaranayake Childrens' Hospital, Peradeniya and District General Hospital, Matale. One hundred faecal samples collected from healthy children were used as controls. All control and test samples were screened for the presence of Cryptosporidium oocysts with Modified Ziehl-Neelsen (MZN) method and PCR. RESULTS: Prevalence of Cryptosporidium infection among children with diarrhoea was 5.7%. All the cases positive for Cryptosporidium were below 3 years of age. The majority (7 out of 8) of the positive cases had watery diarrhoea while none of the healthy children excreted Cryptosporidium oocysts in the faeces. Of the 8 positive cases, 6 had a history of animal contact. A large proportion of positive cases used pipe borne municipal water. The majority (66.6%) of positive cases did not consume boiled cooled water. We were able to identify C. parvum from one of the eight cases that had diarrhoea. CONCLUSIONS: The current study shows that Cryptosporidium is one of the aetiological agents responsible for childhood watery diarrhoea in Sri Lanka, thus stressing the importance of routine stool examination for Cryptosporidium oocysts. This study recommends boiling water as an important measure to prevent the transmission of Cryptosporidium oocysts. Further molecular studies are needed to determine the other species of Cryptosporidium responsible for cryptosporidiosis in children in Sri Lanka.

Demographic and clinical features of suspected dengue and dengue haemorrhagic fever in the Northern Province of Sri Lanka, a region afflicted by an internal conflict for more than 30 years-a retrospective analysis.

OBJECTIVES: The aim of this study was to determine the demographic, clinical, and notification data of suspected dengue fever (DF) and dengue hemorrhagic fever (DHF) cases admitted to Jaffna Teaching Hospital, Sri Lanka. METHODS: The data were collected from bed head tickets of all patients presenting with clinically suspected DF/DHF from October 2009 to September 2010. RESULTS: A total of 1085 clinically suspected DF/DHF cases were identified, with high numbers occurring during December 2009 to March 2010. The majority of the reported patients were females (n = 550, 50.7%) and approximately three-quarters of the patients (n = 797, 73.5%) were adults. All had fever, but fever spikes were noted in only 129 cases (11.9%; 95% confidence interval (CI) 10.1-13.9%). Over 50% of cases had vomiting (95% CI 47.5-53.5%). Haemorrhages were noted in 266 (24.5%), with gum bleeding in 99 patients (37.2%). Low white blood cell and platelet counts were noted in 27.1% and 85.6% of cases, respectively. Of the 1085 cases, only 24 (2.2%) were screened for dengue IgM/IgG and only 458 cases (42.2%) were notified to the Epidemiology Unit, Ministry of Health, Sri Lanka. CONCLUSIONS: The absence of laboratory diagnosis and poor notification to the Epidemiology Unit were the major drawbacks noted.

Evolution of dengue in Sri Lanka-changes in the virus, vector, and climate.

Despite the presence of dengue in Sri Lanka since the early 1960s, dengue has become a major public health issue, with a high morbidity and mortality. Aedes aegypti and Aedes albopictus are the vectors responsible for the transmission of dengue viruses (DENV). The four DENV serotypes (1, 2, 3, and 4) have been co-circulating in Sri Lanka for more than 30 years. The new genotype of DENV-1 has replaced an old genotype, and new clades of DENV-3 genotype III have replaced older clades. The emergence of new clades of DENV-3 in the recent past coincided with an abrupt increase in the number of dengue fever (DF)/dengue hemorrhagic fever (DHF) cases, implicating this serotype in severe epidemics. Climatic factors play a pivotal role in the epidemiological pattern of DF/DHF in terms of the number of cases, severity of illness, shifts in affected age groups, and the expansion of spread from urban to rural areas. There is a regular incidence of DF/DHF throughout the year, with the highest incidence during the rainy months. To reduce the morbidity and mortality associated with DF/DHF, it is important to implement effective vector control programs in the country. The economic impact of DF/DHF results from the expenditure on DF/DHF critical care units in several hospitals and the cost of case management.

Immune response to hepatitis B vaccine in a group of health care workers in Sri Lanka.

Health care workers (HCWs) are considered at high risk of acquiring the hepatitis B virus (HBV). Seroconversion rates after vaccination against HBV among HCWs have not previously been available in Sri Lanka. In the current study, the response to HBV surface antigen (HBsAg) vaccine was assessed in a selected group of HCWs by testing for antibodies against HBsAg (anti-HBs). This was a retrospective descriptive study to measure the anti-HBs levels, using an ELISA, in an immunized group of HCW referred to Department of Microbiology, Faculty of Medicine, University of Peradeniya, Sri Lanka. Among the 342 participants, 9.9% (n=34) were non-responders. Female participants had a significantly higher immune response (94.7%) than males (p<0.05). The results of the study found no significant decline in the immune response with time (p > 0.05). Post HBsAg vaccination immunity in HCW in Sri Lanka is similar to that of global rates with similar gender variation. Anti-HBs levels should be tested in all HCW following HBsAg vaccination so that necessary precautions can be taken.

Infectivity of Cryptosporidium parvum isolated from asymptomatic adult goats to mice and goat kids.

An experimental study was carried out in neonatal goat kids to examine the infectivity of Cryptosporidium oocysts, pattern of oocyst shedding and morphological changes in the intestine during the infection. Cryptosporidium oocysts isolated from adult asymptomatic goats, and identified as C. parvum by polymerase chain reaction (PCR) were used in this study. Of three 4-day-old goat kids, two were orally infected with C. parvum oocysts (10(5) oocysts in 10 ml PBS/kid). One goat kid given 10 ml PBS only by the oral route served as a control. Cryptosporidium oocysts were detected in the faeces of one infected kid on day 3 post-inoculation (pi) whereas in the other 6 days pi. The faecal oocyst counts gradually increased and the peak counts in the two kids were 2 x 10(6)g(-1) (on day 12 pi) and 3.2 x 10(6)g(-1) (on day 14 pi). The increase in faecal oocyst output coincided with diarrhoea in an infected kid from days 10-17 pi. Although the oocyst excretion declined gradually after the peak, both infected kids excreted oocysts until euthanized on days 20 and 22 pi. Light and scanning electron microscopic investigations of the ileum revealed the endogenous stages on the brush border of the enterocytes, infiltration of neutrophils and mononuclear cells into the lamina propria, atrophy, stunting and fusion of villi. For purposes of comparison, goat Cryptosporidium oocysts were inoculated orally (10(3) oocysts/mouse) to eight, 1-week-old mice. All experimental mice excreted oocysts from day 3 pi, and four infected mice continued to excrete oocysts up to day 42 pi. The experimental infection described in goat kids resembled the natural disease in terms of oocyst excretion, clinical signs and intestinal pathology. The ability of oocysts excreted by asymptomatic goats, to infect goat kids and mice is likely to have a major impact on the epidemiology of cryptosporidiosis in livestock and man.

Excretion of Cryptosporidium oocysts by goats in relation to age and season in the dry zone of Sri Lanka.

Pattern of Cryptosporidium oocyst shedding in relation to age and season was investigated monthly from May 1999 to April 2000 in three groups (24 goats per group) of naturally infected goats (from 1 month of age). The three groups designated 1, 2 and 3 were studied for 12, 6 and 3 months, respectively. An association between Cryptosporidium oocyst counts and age was demonstrated. In Group 1, oocyst excretion in the first, second and fourth months of age were significantly higher than that in 6, 7, 8, 9 and 12 months of age (p<0.01), whereas in Group 2, oocyst excretion in the first month of age was significantly higher than that in 2, 4, 5 and 6 months of age (p<0.01). The 3-month observations made in Group 3 showed high oocyst excretion during 1 and 3 months of age. The mean maximum count for Group 1 was 592 oocyst per gram of feces when the animals were 2 months old, while in Groups 2 and 3, this was observed at 3 months of age and the oocyst counts were 787 and 520, respectively. A close association between the prevalence of the Cryptosporidium infection and age of the animal was also observed (p<0.01). At least one-third of the Group 1 animals were excreting Cryptosporidium oocysts during the first 5 months of age. Goats excreted Cryptosporidium oocysts irrespective of the dry or rainy season. The long periods of Cryptosporidium oocyst shedding by goats may have implications for the epidemiology of the disease in susceptible hosts.

Prevalence of Cryptosporidium infection in goats in selected locations in three agroclimatic zones of Sri Lanka.

The prevalence of Cryptosporidium oocysts in the faeces of 1020 goats in three age categories was examined during 1999 in selected locations of three agroclimatic zones of Sri Lanka. The oocysts were demonstrated using the Sheather's sucrose flotation method followed by staining with the modified Ziehl Neelsen technique. Cryptosporidium oocysts were detected in animals from all agroclimatic zones with the highest prevalence of infection in the dry zone (33.6%) compared with 24.7 and 21.7% in the intermediate zones and wet, respectively (P<0.001). Overall, Cryptosporidium oocyst counts were significantly higher in goats of <6 months and 7-12 months of age groups compared with goats of >12 months of age (P<0.001). Cryptosporidium oocysts were detected in 291/1020 (28.5%) animals, while 194/1020 animals (19%), 84/1020 animals (8.2%) and 13/1020 animals (1.3%) excreted low (1-1000 oocysts per gram of faeces), moderate (1000-5000 oocysts per gram of faeces) and high (>5000 oocysts per gram of faeces) counts, respectively. The mean Cryptosporidium count was 383 oocysts per gram of faeces. The majority of the infected goats were asymptomatic. These animals are likely to play an important role in the epidemiology of cryptosporidiosis in goat kids and humans.