Serological Surveillance Research Articles

Insights into measles virus: Serological surveillance and molecular characterization

BackgroundMeasles has been a significant public health concern in Pakistan, especially in the Khyber Pakhtunkhwa (KPK) province, where sporadic and silent epidemics continue to challenge existing control measures. This study aimed to estimate the prevalence and investigate the molecular epidemiology of the measles virus (MeV) in KPK and explore the vaccination status among the suspected individuals. MethodsA cross-sectional study was conducted between February and October 2021. A total of 336 suspected measles cases from the study population were analyzed for IgM antibodies using Enzyme-Linked Immunosorbent Assay (ELISA). Throat swabs were randomly collected from a subset of positive cases for molecular analysis. Phylogenetic analysis of MeV isolates was performed using the neighbor-joining method. The vaccination status of individuals was also recorded. ResultsAmong the suspected participants, 61.0% (205/336) were ELISA positive for IgM antibodies, with a higher prevalence in males (64.17%) compared to females (57.04%). The majority of cases (36.0%) were observed in infants and toddlers, consistent with previous reports. The majority of IgM-positive cases (71.7%) had not received any dose of measles vaccine, highlighting gaps in vaccine coverage and the need for improved immunization programs. Genetic analysis revealed that all MeV isolates belonged to the B3 genotype, with minor genetic variations from previously reported variants in the region. ConclusionThis study provides valuable insights into the genetic epidemiology of the MeV in KPK, Pakistan. The high incidence of measles infection among unvaccinated individuals highlights the urgency of raising awareness about vaccine importance and strengthening routine immunization programs.

Wastewater Surveillance Provides Spatiotemporal SARS-CoV-2 Infection Dynamics

Wastewater surveillance (WWS) can leverage its wide coverage, population-based sampling, and high monitoring frequency to capture citywide pandemic trends independent of clinical surveillance. Here we conducted a nine months daily WWS for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from 12 wastewater treatment plants (WWTPs), covering approximately 80% of the population, to monitor infection dynamics in Hong Kong, China. We found that the SARS-CoV-2 virus concentration in wastewater was correlated with the daily number of reported cases and reached two pandemic peaks three days earlier during the study period. In addition, two different methods were established to estimate the prevalence/incidence rates from wastewater measurements. The estimated results from wastewater were consistent with findings from two independent citywide clinical surveillance programmes (rapid antigen test (RAT) surveillance and serology surveillance), but higher than the cases number reported by the Centre for Health Protection (CHP) of Hong Kong, China. Moreover, the effective reproductive number (Rt) was estimated from wastewater measurements to reflect both citywide and regional transmission dynamics. Our findings demonstrate that large-scale intensive WWS from WWTPs provides cost-effective and timely public health information, especially when the clinical surveillance is inadequate and costly. This approach also provides insights into pandemic dynamics at higher spatiotemporal resolutions, facilitating the formulation of effective control policies and targeted resource allocation.

Development and application of an indirect ELISA for detection of antibodies against emerging atypical porcine pestivirus

BackgroundAtypical porcine pestivirus (APPV) is a newly discovered swine pestivirus, which can cause congenital tremor and high mortality in newborn piglets and subclinical infection in adult pigs, leading to significant impacts on the pig industry. Currently, there is no approved serological method to assess APPV infection status in pig farms.MethodsIn this study, the envelope glycoprotein E2 of APPV was highly expressed in suspension HEK293 cells, and further an indirect enzyme-linked immunosorbent assay based on the recombinant E2 protein (E2-iELISA) was developed and evaluated.ResultsThe reaction parameters of the E2-iELISA were optimized, and the cutoff value was determined to be 0.2 by analyzing S/P values of 165 negative sera against APPV that were confirmed by virus neutralization test (VNT). Specificity test showed that the method had no cross-reaction with other common swine viruses. The E2-iELISA was evaluated using a panel of swine sera, and showed high sensitivity (113/120, 94.2%) and specificity (65/70, 92.9%), and the agreement rate with VNT was 93.7% (178/190). Subsequently, the E2-iELISA was utilized to investigate the seroprevalence of APPV in pig herds of China. When detecting 1368 pig serum samples collected from nine provinces in China, the overall seroprevalence of APPV was 73.9% (1011/1368).ConclusionOur findings suggest that the E2-iELISA is specific and sensitive, and could be a valuable tool for serological surveillance of APPV infection in pigs.

Clinical surveillance systems obscure the true cholera infection burden in an endemic region

Our understanding of cholera transmission and burden largely relies on clinic-based surveillance, which can obscure trends, bias burden estimates and limit the impact of targeted cholera-prevention measures. Serological surveillance provides a complementary approach to monitoring infections, although the link between serologically derived infections and medically attended disease incidence—shaped by immunological, behavioral and clinical factors—remains poorly understood. We unravel this cascade in a cholera-endemic Bangladeshi community by integrating clinic-based surveillance, healthcare-seeking and longitudinal serological data through statistical modeling. Combining the serological trajectories with a reconstructed incidence timeline of symptomatic cholera, we estimated an annual Vibrio cholerae O1 infection incidence rate of 535 per 1,000 population (95% credible interval 514–556), with incidence increasing by age group. Clinic-based surveillance alone underestimated the number of infections and reported cases were not consistently correlated with infection timing. Of the infections, 4 in 3,280 resulted in symptoms, only 1 of which was reported through the surveillance system. These results impart insights into cholera transmission dynamics and burden in the epicenter of the seventh cholera pandemic, where >50% of our study population had an annual V. cholerae O1 infection, and emphasize the potential for a biased view of disease burden and infection risk when depending solely on clinical surveillance data.

Serological surveillance reveals a high exposure to SARS-CoV-2 and altered immune response among COVID-19 unvaccinated Cameroonian individuals.

Surveillance of COVID-19/SARS-CoV-2 dynamics is crucial to understanding natural history and providing insights into the population's exposure risk and specific susceptibilities. This study investigated the seroprevalence of SARS-CoV-2 antibodies, its predictors, and immunological status among unvaccinated patients in Cameroon. A multicentre cross-sectional study was conducted between January and September 2022 in the town of Douala. Patients were consecutively recruited, and data of interest were collected using a questionnaire. Blood samples were collected to determine Immunoglobin titres (IgM and IgG), interferon gamma (IFN- γ) and interleukin-6 (IL-6) by ELISA, and CD4+ cells by flow cytometry. A total of 342 patients aged 41.5 ± 13.9 years were included. Most participants (75.8%) were asymptomatic. The overall crude prevalence of IgM and IgG was 49.1% and 88.9%, respectively. After adjustment, the seroprevalence values were 51% for IgM and 93% for IgM. Ageusia and anosmia have displayed the highest positive predictive values (90.9% and 82.4%) and specificity (98.9% and 98.3%). The predictors of IgM seropositivity were being diabetic (aOR = 0.23, p = 0.01), frequently seeking healthcare (aOR = 1.97, p = 0.03), and diagnosed with ageusia (aOR = 20.63, p = 0.005), whereas those of IgG seropositivity included health facility (aOR = 0.15, p = 0.01), age of 40-50 years (aOR = 8.78, p = 0.01), married (aOR = 0.21, p = 0.02), fever (aOR = 0.08, p = 0.01), and ageusia (aOR = 0.08, p = 0.01). CD4+, IFN-γ, and IL-6 were impaired in seropositive individuals, with a confounding role of socio-demographic factors or comorbidities. Although the WHO declared the end of COVID-19 as a public health emergency, the findings of this study indicate the need for continuous surveillance to adequately control the disease in Cameroon.

Bayesian evaluation of meat juice ELISA for detecting Salmonella in slaughtered pigs without specifying a cut-off.

Consumption of pork and pork products is a major source of human infection with Salmonella. Salmonella is typically subclinical in pigs, making it difficult to identify infected pigs. Therefore, effective surveillance of Salmonella in pigs critically relies on good knowledge on how well the diagnostic tests used perform. A test that has been used in several countries for Salmonella monitoring is serological testing of meat juice using an ELISA (MJ ELISA) to detect antibodies against Salmonella. This MJ ELISA data could be used to estimate infection prevalence and trends. However, as the MJ ELISA output is a sample-to-positive (S/P) ratio, which is a continuous outcome rather than a binary (positive/negative) result, the interpretation of this data depends upon a chosen cut-off. To apply Bayesian latent class models (BLCMs) to estimate diagnostic accuracy of the MJ ELISA test values in the absence of a gold standard without needing to apply a cut-off. BLCMs were fitted to data from a UK abattoir survey carried out in 2006 in order to estimate the diagnostic accuracy of MJ ELISA with respect to the prevalence of active Salmonella infection. This survey consisted of a MJ ELISA applied in parallel with the bacteriological testing of caecal contents, carcass swabs and lymph nodes (n = 625). A BLCM was also fitted to the same data but with dichotomisation of the MJ ELISA results, in order to compare with the model using continuous outcomes. Estimates were obtained for sensitivity and specificity of the ELISA over a range of S/P values and for the bacteriological tests and were found to be similar between the models using continuous and dichotomous ELISA outcomes. The Bayesian method without specifying a cut-off does allow prevalence to be inferred without specifying a cut-off for the ELISA. The study results will be useful for estimating infection prevalence from serological surveillance data.

The Elecsys® Anti-SARS-CoV-2 and Elecsys® Anti-SARS-CoV-2 S antibody assays: Differentiating between vaccination and infection, and assessing long-term performance.

Serological surveillance is useful for assessing SARS-CoV-2 immunity in populations. To effectively study the presence and persistence of antibodies, it is necessary to distinguish between persons with past infection, and persons who only received vaccination. Knowledge of the duration of antibody persistence is essential for correct interpretation of surveillance results. Starting in April 2020, waning of SARS-CoV-2 antibodies was studied in a longitudinal cohort study of 495 SARS-CoV-2 antibody-positive Dutch blood donors, not pre-selected by PCR testing or disease severity. Additionally, in May 2021, a sample of donors representative for the Dutch population was tested for antibodies against the SARS-CoV-2 spike (S) protein, using the Wantai Ab ELISA and the Elecsys® Anti-SARS-CoV-2 S assay; and for antibodies against the nucleocapsid protein, which indicate past infection, using the Elecsys® Anti-SARS-CoV-2 assay. The anti-S response in donors that were infected in April or May 2020 remained positive in 100% of donors in the Elecsys® Anti-SARS-CoV-2 S assay one year after infection, after which follow up of waning was no longer possible because of large scale vaccination. The anti-nucleocapsid response results were still positive in approximately 80% of donors two years after infection. In May 2021, 51% of the donors showed anti-S reactivity and 16.8% tested positive for anti-nucleocapsid antibodies. Infection with SARS-CoV-2 resulted in spike and nucleocapsid antibody levels still detectable in the majority of donors 1-2 years after infection. In May 2021, 51% of donors were vaccinated and 16.8% had had an infection. Thus, both Elecsys® SARS-CoV-2 antibody assays can be used to reliably assess the vaccination and infection status of individuals.

Multiplex Bead Assay for the Serological Surveillance of Measles and Rubella.

There is increasing interest in evaluating antibody responses to multiple antigen targets in a single assay. Immunity to measles and rubella are often evaluated together because immunity is provided through combined vaccines and because routine immunization efforts and surveillance for measles and rubella pathogens are combined in many countries. The multiplex bead assay (MBA) also known as the multiplex immunoassay (MIA) described here combines the measurement of measles- and rubella-specific IgG antibodies in serum quantitatively according to international serum standards and has been successfully utilized in integrated serological surveillance.

Constructing an ELISA for Detection of Anti-Borrelia in Wildlife and Agricultural Animals.

Zoonotic diseases have major impacts on human and animal health, as well as being ecologically significant. Lyme Borreliosis or Lyme disease, caused by infection by pathogenic members of the Borrelia genus, is among these zoonotic diseases. Serology is one of the most accessible means for indirect surveillance of pathogen presence by monitoring the presence, abundance, and type of immune response to the pathogen or pathogen-associated epitopes. Serological surveillance of wild animals is important as wild animals are the primary reservoirs of manyzoonotic diseases. Similarly, serological surveillance of agricultural animals is important due to their economic importance, in addition to animal welfare concerns. However, serology in any non-model animal such as wildlife or agricultural animals is difficult because serology necessarily relies on blood samples from the animals being tested. While companion or laboratory animals are generally sufficiently accustomed to humans that blood samples can be obtained, obtaining blood samples from wild or agricultural animals is more challenging. This initial challenge is compounded by the absence of validated serological tools to evaluate antibody titres in the sera. In this chapter, we provide methods for constructing an ELISA for the detection of anti-Borrelia antibodies in non-model animals, using studies on horses and cows as a proof of principle. The methods focus on the problems specific to non-model animals including obtaining sera, options for determining positive and negative controls without the ability to perform controlled infections, and methods for test optimization and validation.

Serological surveillance of GI norovirus reveals persistence of blockade antibody in a Jidong community-based prospective cohort, 2014-2018.

Herd immunity against norovirus (NoV) is poorly understood in terms of its serological properties and vaccine designs. The precise neutralizing serological features of genotype I (GI) NoV have not been studied. To expand insights on vaccine design and herd immunity of NoVs, seroprevalence and seroincidence of NoV genotypes GI.2, GI.3, and GI.9 were determined using blockade antibodies based on a 5-year longitudinal serosurveillance among 449 residents in Jidong community. Correlation between human histo-blood group antigens (HBGAs) and GI NoV, and dynamic and persistency of antibodies were also analyzed. Seroprevalence of GI.2, GI.3, and GI.9 NoV were 15.1%-18.0%, 35.0%-38.8%, and 17.6%-22.0%; seroincidences were 10.0, 21.0, and 11.0 per 100.0 person-year from 2014 to 2018, respectively. Blockade antibodies positive to GI.2 and GI.3 NoV were significantly associated with HBGA phenotypes, including blood types A, B (excluding GI.3), and O+; Lewis phenotypes Leb+/Ley+ and Lea+b+/Lex+y+; and secretors. The overall decay rate of anti-GI.2 antibody was -5.9%/year (95% CI: -7.1% to -4.8%/year), which was significantly faster than that of GI.3 [-3.6%/year (95% CI: -4.6% to -2.6%/year)] and GI.9 strains [-4.0%/year (95% CI: -4.7% to -3.3%/year)]. The duration of anti-GI.2, GI.3, and GI.9 NoV antibodies estimated by generalized linear model (GLM) was approximately 2.3, 4.2, and 4.8 years, respectively. In conclusion, enhanced community surveillance of GI NoV is needed, and even one-shot vaccine may provide coast-efficient health benefits against GI NoV infection.

Preparation and epitope mapping of monoclonal antibodies against African swine fever virus p22 protein

African swine fever (ASF), caused by the African swine fever virus (ASFV), is now widespread in many countries and severely affects the commercial rearing of swine. Rapid and early diagnosis is crucial for the prevention of ASF. ASFV mature virions comprise the inner envelope protein, p22, making it an excellent candidate for the serological diagnosis and surveillance of ASF. In this study, the prokaryotic-expressed p22 recombinant protein was prepared and purified for immunization in mice. Four monoclonal antibodies (mAbs) were identified using hybridoma cell fusion, clone purification, and immunological assays. The epitopes of mAbs 14G1 and 22D8 were further defined by alanine-scanning mutagenesis. Our results showed that amino acids C39, K40, V41, D42, C45, G48, E49, and C51 directly bound to 14G1, while the key amino acid epitope for 22D8 included K161, Y162, G163, D165, H166, I167, and I168. Homologous and structural analysis revealed that these sites were highly conserved across Asian and European ASFV strains, and the amino acids identified were located on the surface of p22. Thus, our study contributes to a better understanding of the antigenicity of the ASFV p22 protein, and the results could facilitate the prevention and control of ASF.

Laboratory testing and on-site storage are successful at mitigating the risk of release of foot-and-mouth disease virus via production of bull semen in the USA.

Thousands of frozen bovine semen doses are produced daily in the US for domestic use. An incursion of foot-and-mouth disease (FMD) in the country would pose strong challenges to the movements of animals and animal products between premises. Secure supply plans require an estimation of the risk associated with target commodities and the effectiveness of mitigation measures. This study presents the results of a quantitative assessment of the risk of release of FMD virus from five of the largest commercial bull studs in the US via contaminated frozen processed semen. The methodology from a previous study was adapted to better fit the US production system and includes more recent data. Two models were combined, a deterministic compartmental model of FMD transmission and a stochastic risk assessment model. The compartmental model simulated an FMD outbreak within a collection facility, following the introduction of a latent-infected bull. The risk of release was defined as the annual likelihood of releasing at least one frozen semen batch, defined as the total amount of semen collected from a single bull on a given collection day, containing viable FMD virus. A scenario tree was built using nine steps leading from the collection to the release of a contaminated batch from a given facility. The first step, the annual probability of an FMD outbreak in a given facility, was modeled using an empirical distribution fitted to incidence data predicted by five models published between 2012 and 2022. An extra step was added to the previously published risk pathway, to account for routine serological or virological surveillance within facilities. The results showed that the mitigation measures included in the assessment were effective at reducing the risk of release. The median annual risk of release from the five facilities was estimated at less than 2 in 10 billion (1.5 x 10-10) in the scenario including a 30-day storage, routine genome detection assays performed every two weeks and RT-PCR testing of the semen. In this scenario, there was a 95% chance that the risk of release would be lower than 0.00041. This work provides strong support to the industry for improving their response plans to an incursion of FMD virus in the US.

Spatiotemporal description of African swine fever virus nucleic acid and antibodies detected in pigs sampled at abattoirs in the greater Kampala metropolitan area, Uganda from May 2021 through June 2022

BackgroundAfrican swine fever virus (ASFV) infections in Africa cause hemorrhagic disease in domestic pigs and is maintained by a sylvatic cycle in warthogs. It is endemic in Uganda, leading to significant economic losses. Previous studies performed in rural areas and in Kampala had differing diagnostic results. The purpose of this study was to provide a robust spatial, temporal, and diagnostic summary of pigs slaughtered in the greater Kampala metropolitan area over the course of one year. This study characterized 1208 to 1323 serum, blood, and tissue samples collected from pigs at six abattoirs in the greater Kampala metropolitan area of Uganda monthly from May 2021 through June 2022. Validated and standardized serologic and molecular diagnostics were used.ResultsOnly 0.15% of pigs had detectable antibodies against ASFV, suggesting low survival rates or pre-clinical diagnosis. Yet, 59.5% of pigs were positive for ASFV DNA. Blood had the lowest detection rate (15.3%) while tonsil and lymph nodes had the highest (38% and 37.5%, respectively), spleen samples (31.5%) were in between. Agreement between sample types was fair to moderate overall. A significant seasonality of ASFV infections emerged with infections found predominately in the dry seasons. Spatial assessments revealed that the greater Kampala metropolitan area abattoirs have a catchment area that overlaps with Uganda’s most pig dense regions.ConclusionsPigs at greater Kampala metropolitan area abattoirs can be sentinels for acute disease throughout the pig dense region of Uganda, particularly in the dry seasons. The high prevalence detected suggests that pigs are sold in response to local reports of ASFV infections (panic sales). Serological surveillance is not useful, as very few pigs seroconverted in this study prior to slaughter. In contrast, tissue samples of pigs can be used to detect disease using qPCR methods.

Avian reovirus: a furious and fast evolving pathogen.

Avian reoviruses (ARVs) have a significant economic impact on the poultry industry, affecting commercial and backyard flocks. Spread feco-orally, or vertically, many do not cause morbidity, but pathogenic strains can contribute to several diseases, including tenosynovitis/arthritis, which is clinically the most significant. The last decade has seen a surge in cases in the US, and due to ongoing evolution, seven genotypic clusters have now been identified. Control efforts include strict biosecurity and vaccination with commercial and autogenous vaccines. Research priorities include improving understanding of pathogenesis and developing new vaccines guided by ongoing molecular and serologic surveillance.

Knowledge and perceptions on the existence of Voluntary Counseling and Testing (VCT) Centers and associated factors among Brazilian LGBT+ individuals.

In Brazil, the advent of voluntary and counseling testing (VCT) has provided many benefits in the fight against AIDS. A sectional open web survey was conducted to investigate the Brazilian LGBT+ individuals' knowledge and perceptions of the existence of VCT centers and their associations with risk behavior and other variables of interest. The study instrument included questions on sociodemographic and behavioral variables, knowledge on VCT, access to and use of it, beliefs about HIV infection, test results, and risk perception. The Risk Behavior Score for HIV infection (RBS) was also used. A total of 1,630 volunteers participated in the survey, of whom 56.8% were men. Of these, 96.1% declared themselves as (MSM). Almost 50% of the participants had never heard about the existence of VCT, which was a higher knowledge score among MSM. A low frequency of participants had a clear perception of the risks to which they are exposed, whereas those with higher RBS were aware of the existence of VCT. As VCT is critical for the implementation of policies to combat and prevent HIV and other sexually transmitted diseases, our results can contribute to a better understanding of the influence of VCT on the frequency of testing, serological surveillance, and routine counseling for key populations.

Estimating prevalence of Enterovirus D111 in human and non-human primate populations using cross-sectional serology.

Enteroviruses primarily affect young children with a varying severity of disease. Recent outbreaks of severe respiratory and neurological disease due to EV-D68 and EV-A71, as well as atypical hand-foot-and-mouth-disease due to CVA6, have brought to light the potency of enteroviruses to emerge as severe human pathogens. Enterovirus D111 (EV-D111) is an enteric pathogen initially detected in Central Africa in human and wildlife samples and was recently detected in environmental samples. The natural history and epidemiology of EV-D111 are poorly studied. Here, the presence of serum neutralizing antibodies to EV-D111 was estimated in human and wildlife samples from five countries. We report high prevalence of neutralizing antibodies measured against EV-D111 in human populations (range, 55-83 %), a proxy for previous infection, which indicates active virus circulation in absence of detection in clinical cases and a high number of undiagnosed infections. Notably, seroprevalence in samples from the UK varied by age and was higher in children and older adults (1-5 and >60 years old), but lower in ages 11-60. EV-D111 seroprevalence in apes and Old World monkeys was 50 % (33-66 %), which also suggests prior exposure and supports existing knowledge of enterovirus circulation in wild and captive apes and Old World monkeys. Generally, reported cases of infection likely underestimate the prevalence of infection particularly when the knowledge of community transmission is limited. Continued serologic surveillance and detection of EV-D111 in clinical and environmental samples will allow for a more robust assessment of EV-D111 epidemiology.

HPAI H5N1 outbreak in Chile, 2022-2023. Proceeding of The First International Avian Influenza Summit, University of Arkansas- October 16-17, 2023"

In Chile, from December 5, 2022, to date (August 30), 892 HPAI H5N1 HPAI-positive samples have been detected in wild birds, affecting 50 species, with 94 zones under control. To date, more than 35,000 wild birds are dead. In addition, 175 positive backyard bird premises have been detected, with more the 14,000 birds culling distributed in the 16 regions of the country in 97 zones under sanitary control. This is the event 4,775 registered in the WAHIS system. In addition, since March 13, 2023, the HPAI H5N1 virus was identified in a broiler genetics unit. Since that date, 12 outbreaks have been detected nationwide, which were distributed in 12 flocks (1.47\% of the total number of industrial flocks). Of these, 5 were in laying hens (1.4\% of the national total), 3 in broiler turkeys, 2 in turkey breeders, 1 in broilers, and 1 in broiler breeders. As a result of these outbreaks, the total number of susceptible birds was eliminated, corresponding to 1,479,433 birds. The procedure for each poultry outbreak is determined in the current AI Contingency Plan, which defines in general terms the establishment of a Zone under Sanitary Control (ZBCS), where a protection zone (3 km radius) and a surveillance zone of 7 km radius are defined, forming a total restriction zone of 10 km radius around each outbreak. For outbreaks in poultry, 7 ZBCS were created, concentrating all commercial outbreaks. commercial outbreaks. In some cases (as in ZBCS\_102 and 114), more than one outbreak was detected in each one. The actions carried out within each ZBCS in wildlife and in poultry and non-poultry are surveillance in the protection and surveillance zones, including clinical, molecular, and serological surveillance. Chile declared itself free of highly pathogenic avian influenza on August 22, 2023. On the other hand, on February 16, 2023, the first case of HPAI H5N1 in aquatic mammals was recorded, being a sea lion, the affected individual. To date, more than 20,000 sea lions have been stranded and died from the disease, and cases have been reported in other aquatic mammal species. In this outbreak, one human case has been reported in the north of the country and one terrestrial mammal (Geoffroy’s cat) in Patagonia. The Chilean A/H5 HA genes were genetically closest related to samples detected in Peru around the same period, and both Peruvian and Chilean strains derive from A/H5N1 viruses, causing widespread outbreaks in poultry and wild birds across North America. This outbreak corresponds to the first incursion of a HPAI H5 virus of Euro-North American lineage to South America, having serious consequences for wildlife, affecting backyard poultry and commercial flocks, which forces us to improve our capacities for risk assessment, surveillance, biosecurity, diagnosis, training and risk communication to poultry farmers, professionals and publishers in Chile as well as in Latin America.

Experiences in Peru for surveillance in the fight against highly pathogenic H5N1 avian influenza. Proceeding of The First International Avian Influenza Summit, University of Arkansas- October 16-17, 2023"

The emergence of highly pathogenic H5N1 avian influenza poses a significant threat to human and animal health worldwide. This summary provides an overview of the surveillance efforts implemented in Peru to combat the spread of H5N1 avian influenza. Joint efforts between the authority, the productive sector, and academia date back to the first influenza outbreak in Chile in 2002. Since then, the government has been developing intense clinical and serological surveillance in the domestic birds and poultry industry. In parallel, the university has been carrying out important research to identify and typify the viruses that circulate in the resident and migratory wild bird populations of the Peruvian coast, having detected and identified during these years around 50 strains of influenza viruses of 10 different hemagglutinin subtypes and 8 neuraminidases, a single strain of the low pathogenicity H7N3 subtype. Until then, an H5-type virus had never been detected in wild birds. With the arrival of the virus in North America, a comprehensive response was developed in which the key components of the surveillance program included active and passive clinical surveillance in wild bird populations, commercial poultry, and backyard and game bird populations. Regarding surveillance of wild birds, the authority has received 238 notifications, of which 64 were confirmed positive, from 15 regions. In the Avian Pathology Laboratory of the UNMSM, actions were carried out to support the active surveillance of wild birds of the Peruvian coast in the impact zones for poultry farming, analyzing 405 samples of fresh feces from 41 species of asymptomatic coastal aquatic wild birds, being positive 12 (2.96\%). Additionally, 45 samples of wild birds with suspicious symptoms were analyzed and sent by different government institutions (SENASA, SERFOR, SERNAMP), with 19 (42\%) being confirmed positive. Regarding domestic bird populations, 1,676 notifications have been made to SENASA, of which 194 were positive for the H5N1 virus in 13 country regions. On the other hand, the Avian Pathology Lab received 16 confirmed positive cases by PCR and viral isolation in SPF embryonated eggs from ducks, quails, layers, native birds, and one case of turkeys. The complete genome analysis of 09 strains of wild birds, 04 of domestic birds, and one of sea lions has been carried out. Analysis of new samples is in process. NGS analysis has identified the IA H5N1 clade 2.3.4.4b virus; however, it has been found that the Peruvian strains are lining up in a group together with the Chilean strains. All these activities are possible to carry out thanks to the financial support of national and international institutions: NAMRU, Concytec-World Bank, Peruvian Poultry Association, and ALA-US Poultry. The high vulnerability of poultry populations in the country is due to its location along the coast, where tens of thousands of dead wild birds have fallen, in addition to being surrounded by a very high population of ducks, fighting, and backyard birds. The government determined emergency vaccination, as well as the continuation of clinical and virological surveillance in vaccinated and unvaccinated poultry populations. SENASA has been evaluating the protection of approved vaccines, both by the serological response with hemagglutination inhibition tests using local antigens and by challenge. The industry has responded by significantly increasing the levels of structural and operational biosafety, as well as directly supporting control actions at the regional level. Since March, no outbreaks have been reported in industrially farmed birds.

Epidemiological features of Leishmania infantum in dogs (Canis lupus familiaris) suggest a latent risk of visceral leishmaniasis in the metropolitan area of Bucaramanga, Santander, Eastern Colombia.

Visceral leishmaniasis (VL) is a disease caused by species of the Leishmania donovani complex that is mainly transmitted through the urban cycle involving dogs as the primary reservoir. In Colombia, the incidence of VL is increasing, along with the spread of potential vectors. This study aims to investigate the eco-epidemiological factors associated with Leishmania spp. infection in dogs from the Metropolitan Area of Bucaramanga (MAB), Santander, eastern Colombia, which is a region at risk for VL. We conducted molecular and serological surveillance of Leishmania spp. in 207 dogs from MAB to determine the epidemiological factors associated with infection. Subsequently, we carried out a molecular and serological analysis of phlebotomine and humans, respectively, in areas with a higher prevalence of infection, aiming to describe the main features associated with the transmission cycle. Out of the 207 dogs tested, 37 (17.8%, 95% CI=12.6-23.1%) were positive for the presence of Leishmania antibodies by the IFAT test, and only 9 (4.3%, 95% CI=1.55-7.15%) were positive for L. infantum by PCR. Multivariate analyses indicated that canine shelters and dogs with clinical signs commonly associated with canine VL had a higher prevalence of infection (P<0.05). In the entomological survey, 69 blood-fed female phlebotomine of the genus Lutzomyia were captured in canine shelters, among them, 55% were identified as Lutzomyia camposi, 29% as Lu. ovallesi, 7% as Lu. dubitans, 6% as Lu. torvida, and 3% as Lu. cayennensis. The identified meal sources of the phlebotomine included human, pig, avian, cattle, and porcupine (Coendou quichua) blood. However, no phlebotomine positive for Leishmania spp. were detected by molecular analyses. Finally, 14 humans who had frequent contact with L. infantum-positive dogs were analyzed through rK39 test, but none tested was positive for IgG/IgM antibodies. The molecular and serological analyses indicate the circulation of L. infantum in dogs from MAB, with canine shelters having the highest prevalence of infection. The entomological survey of canine shelters showed a significant diversity of phlebotomine without potential vectors of L. infantum, suggesting the presence of infection in dogs from these areas could take place in other locations or through other transmission routes. The circulation of L. infantum in multiple dogs from MAB suggests a latent risk of zoonotic transmission of VL in these cities.

Environmental Surveillance for Salmonella Typhi and its Association With Typhoid Fever Incidence in India and Malawi.

Environmental surveillance (ES) for Salmonella Typhi potentially offers a low-cost tool to identify communities with a high burden of typhoid fever. We developed standardized protocols for typhoid ES, including sampling site selection, validation, characterization; grab or trap sample collection, concentration; and quantitative PCR targeting Salmonella genes (ttr, staG, and tviB) and a marker of human fecal contamination (HF183). ES was implemented over 12 months in a historically high typhoid fever incidence setting (Vellore, India) and a lower incidence setting (Blantyre, Malawi) during 2021-2022. S. Typhi prevalence in ES samples was higher in Vellore compared with Blantyre; 39/520 (7.5%; 95% confidence interval [CI], 4.4%-12.4%) vs 11/533 (2.1%; 95% CI, 1.1%-4.0%) in grab and 79/517 (15.3%; 95% CI, 9.8%-23.0%) vs 23/594 (3.9%; 95% CI, 1.9%-7.9%) in trap samples. Detection was clustered by ES site and correlated with site catchment population in Vellore but not Blantyre. Incidence of culture-confirmed typhoid in local hospitals was low during the study and zero some months in Vellore despite S. Typhi detection in ES. ES describes the prevalence and distribution of S. Typhi even in the absence of typhoid cases and could inform vaccine introduction. Expanded implementation and comparison with clinical and serological surveillance will further establish its public health utility.