Investigation of an outbreak caused by an unknown infectious pathogen in rural Zola.

This case study was written as part of a fellowship in biosafety and biosecurity organised by the German Biosecurity Programme, namely the Global Partnership Initiated Biosecurity Academia for Controlling Health Threats (GIBACHT). Among other objectives, the fellowship focuses on equipping participants with the skills of developing their own country-specific case studies with focus on biosafety- and biosecurity-related scenarios. Upon completion of the underlying case study, participants should be able to identify some existing gaps with regards to early detection and investigation of outbreaks, describe the key steps in outbreak investigation, explain the role of communication and coordination among the various stakeholders in outbreak investigations and analyse epidemiological data obtained during outbreak investigations. They should also be able to suggest appropriate control and prevention measures for specific disease outbreaks with focus on foodborne outbreaks and to distinguish between biosafety and biosecurity concepts.

Containment of a fatal and highly infectious disease outbreak.

This case study was designed based on several experience with several viral haemorrhagic fevers (VHFs) outbreaks responded to in Uganda between 2000 and 2016. Fictitious scenarios have been included to facilitate learning of the users. The major goal of the case study is to facilitate learners to appreciate incident detection and the incident management processes for control and containment of a fatal and highly infectious viral disease outbreak. This case study is targeted towards health scientists of medicine, nursing, biomedical laboratory and public health background. We specifically orient learners on clinical presentation of viral infections and laboratory tests considered for incident detection, conducting a risk assessment for an infectious disease, Infection Prevention and Control in the outbreak setting, skills of incident management, analysis and interpretation of epidemiological data to aid epidemic response and control decisions.

Cholera outbreak in Funpur, Winland.

Vibrio Cholerae is a category B agent which has moderate to high potential to be used in bioterrorist events. This fictitious case study is based on man-made outbreak investigation and response carried out by disease surveillance and response unit of country Winland. The numbers of acute watery diarrhoea cases (AWDs) were concentrated in city Funpur of country Winland which share international border with Robiland, another country with poor health infrastructure. Regular movement of nomadic population between two countries has additional risk of international spread. This case study is designed for the training of public health students and workers on steps of outbreak investigation, packaging of biological samples, understanding IHR reporting algorithm, understanding difference between biosafety and biosecurity, different categories of bioterrorism organisms and PPE & its zones. This case study can be used as supporting training tool for application of learned concepts to a real situation and can be carried out in 2-3 hours.

Outbreak investigation of an unknown gastrointestinal illness in District Victoria, Country Mala, 2016.

This case study is based on an outbreak investigation conducted by multisectoral team from animal and public health offices in Kaktong (a remote village in Zhemgang District Bhutan) during July-September 2010. This outbreak caused by ingestion of infected cow meat which had died after a brief illness (bleeding of unclotted blood from nostrils). The owner of the affected cow had opened the carcass and dressed the meat, which he shared or sold within the village for human consumption. It simulates an epidemiological investigation including active and passive case finding, descriptive and analytical epidemiology, laboratory confirmation, risk communication with implementation of control measures. This case study is designed for the training of front-line public health professional, basic, intermediate and advanced level field epidemiology trainees. The case study will build the capacity of the trainees regarding investigating illnesses caused by animal-human interface.

Emergence of a febrile illness of unknown causes among the population and visitors of Upper Egypt.

This is a fictional case study for training that encourages participants to interact and apply theory into practice. A febrile illness of unknown cause that occurred in Upper Egypt in 2002 was chosen for the events. The location of Aswan was selected to define the climate, topography, and location with the characteristics that support the events. Data obtained from applied research work in Egypt was included. The case study deals with the incidence of severe cases of fever of unknown origin accompanied by neurological and intestinal symptoms, as well as a high percentage of deaths. Most of the symptoms appear in people with direct contact with farm animals especially equines and birds, or those who were near waterways, either tourism workers or tourists. Most of the infected cases or deaths have accumulated in Aswan and some in the neighbouring governorates. This case study focusses on the steps taken during an outbreak investigation, and deals with investigative challenges as well as concepts of biosafety and biosecurity.

Exposure of domestic swine to influenza A viruses in Ghana suggests unidirectional, reverse zoonotic transmission at the human-animal interface.

Influenza A viruses (IAVs) have both zoonotic and anthroponotic potential and are of public and veterinary importance. Swine are intermediate hosts and 'mixing vessels' for generating reassortants, progenies of which may harbour pandemic propensity. Swine handlers are at the highest risk of becoming infected with IAVs from swine but there is little information on the ecology of IAVs at the human-animal interface in Africa. We analysed and characterized nasal and throat swabs from swine and farmers respectively, for IAVs using RT-qPCR, from swine farms in the Ashanti region, Ghana. Sera were also analysed for IAVs antibodies and serotyped using ELISA and HI assays. IAV was detected in 1.4% (n = 17/1,200) and 2.0% (n = 2/99) of swine and farmers samples, respectively. Viral subtypes H3N2 and H1N1pdm09 were found in human samples. All virus-positive swine samples were subtyped as H1N1pdm09 phylogenetically clustering closely with H1N1pdm09 that circulated among humans during the study period. Phenotypic markers that confer sensitivity to Oseltamivir were found. Serological prevalence of IAVs in swine and farmers by ELISA was 3.2% (n = 38/1,200) and 18.2% (n = 18/99), respectively. Human H1N1pdm09 and H3N2 antibodies were found in both swine and farmers sera. Indigenous swine influenza A viruses and/or antibodies were not detected in swine or farmers samples. Majority (98%, n = 147/150) of farmers reported of not wearing surgical mask and few (4%, n = 6) reported to wear gloves when working. Most (n = 74, 87.7%) farmers reported of working on the farm when experiencing influenza-like illness. Poor husbandry and biosafety practices of farmers could facilitate virus transmission across the human-swine interface. Farmers should be educated on the importance of good farm practices to mitigate influenza transmission at the human-animal interface.

Understanding attitude, practices and knowledge of zoonotic infectious disease risks among poultry farmers in Ghana.

Zoonotic infectious diseases (ZIDs) are increasing globally, and livestock farmers in low- and middle-income countries are at particularly high risk. An evaluation of farmer's behaviour on farms can be used to identify the risk factors and to develop tailored control strategies. This study documents the knowledge of zoonotic poultry diseases (ZPD) among 152 poultry farm workers (respondents) from 76 farms in the Ashanti region of Ghana and assessed their on-farm attitude and practices that increase their risk to exposure of ZPD. The median age of respondents was 29 years, 91.4% (n = 139) had a formal education, and 80.9% (n = 123) had worked on the farm for more than 1 year. The majority of farms (n = 69, 90.8%) had multiple flocks and 27.6% (n = 21) kept other animals, of which 57.1% (n = 12) were pigs. The majority of respondents had good knowledge about poultry diseases but not about ZPD. A higher level of education and longer work experience improved respondents' knowledge of poultry and ZPD. Although respondents identified the wearing of personal protective equipment (PPE) as a major ZPD preventive measure, the majority did not put that knowledge into practice. Most farms (71.1%, n = 54) had no footbath and 55.3% (n = 42) deposited farm-waste on the farm. While 97.4% (n = 148) of respondents washed their hands after working, only 48.7% (n = 74) wore protective footwear, 2.7% (n = 4) wore overalls, 2% (n = 3) wore nose masks and none (n = 0) wore gloves. The husbandry practices and attitude of farmers expose them to pathogens on the farm and increase their risk of becoming infected with ZPD in the sub-region. The results from this study could be used to promote human health among farm workers in Ghana.

Fluoroquinolone-Resistant Salmonella enterica, Campylobacter spp., and Arcobacter butzleri from Local and Imported Poultry Meat in Kumasi, Ghana.

Salmonella and Campylobacter are important gastroenteric pathogens. Arcobacter butzleri is an emerging enteric pathogen. Data on the frequencies of these poultry-associated pathogens on meat products sold in sub-Saharan Africa are scarce. This study aimed to analyze the frequency of Salmonella, Campylobacter, and Arcobacter antibiotic resistance and underlying mechanisms of resistance to fluoroquinolones in locally produced and imported poultry sold in urban Ghana. Chicken meat was collected and cultured on standard media. Bacterial strains were identified by biochemical methods and by mass spectrometry. Antibiotic susceptibility was tested by disk diffusion. Ciprofloxacin-resistant strains were assessed for molecular mechanisms of resistance. Among 200 samples, comprising 34% (n = 68) from the Ghanaian poultry industry and 66% (n = 132) from imports, 9% (n = 17) contained Salmonella, 11% (n = 22) Campylobacter, and 26.5% (n = 53) A. butzleri. Higher overall contamination frequencies were found in local meat. Most common Salmonella serovars identified were Kentucky (n/N = 5/16; 31%) and Poona (n/N = 4/16; 25%). Campylobacter were C. coli (n/N = 10/19; 53%) and C. jejuni (n/N = 9/19; 47%). Resistance to fluoroquinolones was high with 63% (n = 10), 75% (n = 15), and 52% (n = 25) in Salmonella, Campylobacter, and Arcobacter, respectively. A link between Salmonella Kentucky [sequence type (ST) 198] and a ciprofloxacin minimum inhibitory concentration of 16 µg/mL was found. Salmonella Poona-ST308 revealed transferable qnrB2 fluoroquinolone resistance genes. Markedly high frequencies of resistant Salmonella, Campylobacter, and Arcobacter predominant in locally produced meat represent a probable transmission reservoir for human infections. These findings highlight the need for implementation of surveillance systems that focus on food hygiene, use of antibiotics in animal husbandry, and continuous monitoring of the quality of meat products from imports.

Equinus Correction During Multilevel Surgery in Adults With Cerebral Palsy.

BACKGROUND: Equinus foot deformity constitutes a common gait disorder in ambulatory adults with bilateral spastic cerebral palsy (BSCP). The outcome after intramuscular aponeurotic lengthening in the context of single-event multilevel surgery (SEMLS) in adulthood has not been investigated. METHODS: We followed a group of 31 ambulatory adults with BSCP and equinus who underwent SEMLS including gastrocnemius-soleus intramuscular aponeurotic recession or Achilles tendon lengthening. All patients were analyzed preoperatively and at least 1 year (mean follow-up period: 1.6 years) postoperatively by clinical examination and 3-dimensional instrumented gait analysis including the Gait Profile Score (GPS). RESULTS: Clinical examination showed no significant improvement of ankle dorsiflexion ( P = .5) and an unchanged plantarflexion ( P = .7) with knee extended but a significant postoperative reduction of spasticity in the calf muscle ( P = .0001) as measured by clinical examination following the modified Ashworth scale. Significant improvement of mean ankle dorsiflexion in stance and swing ( P = .0001) was found. The GPS decreased and improved significantly (15.9 ± 4.6 to 11.4 ± 3.1; P = .0001). Persistence of equinus and calcaneal gait indicating under- and overcorrection at follow-up was found in 1 patient (3%), respectively. CONCLUSION: Intramuscular gastrocnemius-soleus aponeurotic recession is part of multilevel surgery corrected equinus deformity in adults. The increase in muscle length led to significant improvement of kinetic and kinematic parameters during walking without a loss of muscle strength and push-off capacity. The risk of overcorrection after equinus correction in adults with BSCP was found to be relatively low. LEVEL OF EVIDENCE: Level IV, retrospective case series.

No serological evidence for Zika virus infection and low specificity for anti-Zika virus ELISA in malaria positive individuals among pregnant women from Madagascar in 2010.

It was previously reported that a malaria infection may interfere with the specificity of a commercial ELISA test against Zika virus (ZIKV). We analyzed 1,216 plasma samples from healthy, pregnant women collected in two sites in Madagascar in 2010 for ZIKV antibodies using a commercial ELISA and for Plasmodium infection by PCR. This screen revealed six putative ZIKV-positive samples by ELISA. These results could not be confirmed by indirect immunofluorescence assays or virus neutralization tests. Four of these six samples were also positive for P. falciparum. We noted that the frequency of malaria positivity was higher in ZIKV-ELISA positive samples (50% and 100% in the two study sites) than ZIKV-negative samples (17% and 10%, respectively), suggesting that malaria may have led to false ZIKV-ELISA positives.

Antibiotic resistance and clonal diversity of invasive Staphylococcus aureus in the rural Ashanti Region, Ghana.

BACKGROUND: Staphylococcus aureus is among the most common pathogens isolated from blood cultures in Ghana; yet the epidemiology of blood infections in rural settings is poorly described. This study aims to investigate antimicrobial susceptibility and clonal diversity of S. aureus causing bloodstream infections in two hospitals in the Ashanti Region, Ghana. METHODS: Blood cultures were performed for all febrile patients (≥37.5 °C) on hospital admission. Antibiotic susceptibility testing for S. aureus isolates was carried out by the VITEK 2 system. Multiplex polymerase chain reaction (PCR) was used to detect S. aureus-specific nuc gene, Panton-Valentine leukocidin (PVL), and methicillin-resistant S. aureus (MRSA)-specific mecA and mecC genes. The population structure of S. aureus was assessed by spa typing. RESULTS: In total, 9,834 blood samples were cultured, out of which 0.6% (n = 56) were positive for S. aureus. Multidrug resistance (MDR) was detected in 35.7% (n = 20) of the S. aureus strains, of which one was a MRSA. The highest rate of antibiotic resistance was seen for commonly available antibiotics, including penicillin (n = 55; 98.2%), tetracycline (n = 32; 57.1%) and trimethoprim/sulfamethoxazole (n = 26; 46.4%). Of all S. aureus strains, 75.0% (n = 42) carried the PVL-encoding genes. We found 25 different spa types with t355 (n = 11; 19.6%), t314 (n = 8; 14.3%), t084 (n = 8; 14.3%) and t311 (n = 5; 8.9%) being predominant. CONCLUSION: The study exhibited an alarmingly large level of antibiotic resistance to locally available antibiotics. The frequency of genetically diverse and PVL-positive methicillin-sensitive S. aureus (MSSA) was high and could represent a reservoir for the emergence of virulent PVL-positive MRSA clones.

Complete Genome Sequence of a Reassortant H14N2 Avian Influenza Virus from California.

We report the complete genome sequence of a reassortant H14N2 avian influenza virus isolated in 2011 from a northern shoveler in California. This introduced Eurasian subtype acquired seven segments from North American viruses and circulated in the Pacific Flyway 1 year after its detection in the Mississippi Flyway.

Evaluation of phenotypic markers in full genome sequences of avian influenza isolates from California.

We evaluated phenotypic markers in full-genome sequences of avian influenza isolates to identify avian strains with increased potential for transmission and pathogenicity in mammals. Of 149 markers examined, 67 were positive in the consensus sequences from 206 avian isolates. Analysis of deep sequencing data in a subset of 24 isolates revealed that 344 subpopulations occurred at marker positions. Markers in subpopulations were significantly more likely to be negative (258/344) than positive (86/344), but nearly all of the marker-positive subpopulations (78/86) were associated with marker-negative consensus sequences. Our analysis revealed significant variation in important markers among avian isolates, and showed that consensus sequences do not fully convey an isolate's potential for increased transmissibility and pathogenicity in mammals.

Zona pellucida birefringence correlates with developmental capacity of bovine oocytes classified by maturational environment, COC morphology and G6PDH activity.

In the present study we aimed to analyse structural changes during in vitro maturation of the bovine zona pellucida (ZP) by scanning electron microscopy (SEM) ands zona pellucida birefringence (ZPB). Here we show that alterations during in vitro maturation invasively analysed by SEM are reflected in ZPB. In vivo-matured oocytes displayed significantly lower birefringence parameters and significantly higher blastocyst rates compared with in vitro-derived oocytes (39.1% vs 21.6%). The same was observed for in vitro-matured oocytes with cumulus-oocyte complex (COC) Quality 1 (Q1) compared with Q3-COCs with respect to zona birefringence and developmental capacity. Immature oocytes with Q1-COCs displayed higher ZPB values and a higher developmental capacity to the blastocyst stage (27.7% vs 16.9%) compared with immature Q3-COCs. Considering in vitro-matured oocytes, only those with Q1-COC showed a trend for ZPB similar to in vivo-matured oocytes. Therefore, a decreasing trend for ZPB during in vitro maturation seems to be typical for high-quality oocytes and successful cytoplasmic maturation. In accordance, fully-grown immature oocytes reached significantly higher blastocyst rates (32.0% vs 11.5%) and lower ZPB values compared with still-growing ones. In conclusion, we successfully evaluated the applicability of zona imaging to bovine oocytes: alterations during in vitro maturation invasively analysed by scanning electron microscopy were reflected in the birefringence of the zona pellucida of bovine oocytes affecting developmental capacity at the same value. Therefore ZPB measurement by live zona imaging has potential to become a new tool to assess correctness of in vitro maturation and to predict developmental competence.

West Nile virus and its emergence in the United States of America.

Zoonotic West Nile virus (WNV) circulates in natural transmission cycles involving certain mosquitoes and birds, horses, humans, and a range of other vertebrates are incidental hosts. Clinical infections in humans can range in severity from uncomplicated WNV fever to fatal meningoencephalitis. Since its introduction to the Western Hemisphere in 1999, WNV had spread across North America, Central and South America and the Caribbean, although the vast majority of severe human cases have occurred in the United States of America (USA) and Canada. By 2002-2003, the WNV outbreaks have involved thousands of patients causing severe neurologic disease (meningoencephalitis and poliomyelitis-like syndrome) and hundreds of associated fatalities in USA. The purpose of this review is to present recent information on the epidemiology and pathogenicity of WNV since its emergence in North America.

Viral determinants in the NS3 helicase and 2K peptide that promote West Nile virus resistance to antiviral action of 2',5'-oligoadenylate synthetase 1b.

The interferon-inducible 2',5'-oligoadenylate synthetase 1b (Oas1b) protein inhibits West Nile virus (WNV) infection by preventing viral RNA (vRNA) accumulation in infected cells. Serial passage of WNV in Oas1b-expressing mouse cells selected a virus variant with improved growth capacity. Two major amino acid substitutions were identified in this Oas1b-resistant WNV variant: NS3-S365G in the ATPase/helicase domain of NS3 and 2K-V9M in the C-terminal segment of NS4A. To assess their effect on antiviral activity of Oas1b, the NS3 and 2K mutations were engineered into an infectious WNV cDNA clone. The NS3 mutation alters requirement of ATP for ATPase activity and attenuates Oas1b-mediated suppression of vRNA accumulation. However, growth of NS3-mutant virus remains impaired in Oas1b-expressing cells. Only the 2K-V9M mutation efficiently rescued viral growth by promoting vRNA replication. Thus, WNV resistance to Oas1b antiviral action could be attributed to the 2K-V9M substitution with a potential role of NS3-S365G through rescue of vRNA accumulation.

Human genetic determinants of dengue virus susceptibility.

Dengue virus (DENV) is an emerging mosquito-borne pathogen that produces significant morbidity worldwide resulting in an estimated 50-100 million infections annually. DENV causes a spectrum of illness ranging from inapparent infection to life-threatening hemorrhagic fever and shock. The varied DENV disease outcome is determined by complex interactions between immunopathologic, viral, and human genetic factors. This review summarizes these interactions with a focus on human genetic determinants of DENV susceptibility, including human leukocyte antigens, blood type, and single nucleotide polymorphisms in immune response genes that have been associated with DENV disease. We also discuss other factors related to DENV outcome including viral genetic determinants, age, ethnicity, and nutritional status as they relate to DENV susceptibility. We emphasize the need for functional genetics studies to complement association-based data and we call for controlled study designs and standard clinical DENV disease definitions that will strengthen conclusions based on human genetic DENV studies.

Hepatitis C virus-induced secretion of inflammatory chemokines preferentially recruits NKG2A+CD8+ T cells.

In patients with hepatitis C, a loss-of-function mutation of chemokine receptor CCR5 (CCR5Delta32) has been shown to be associated with spontaneous viral clearance and lower levels of hepatic inflammation. In the present study, we show that CCR5 is coexpressed with the inhibitory NKG2A receptor on CD8(+) T cells. Consequently, CCR5(+) T cells were highly susceptible to NKG2A-mediated inhibition of cytotoxic activity and NKG2A(+) lymphocytes were preferentially attracted by CCR5 ligands induced by hepatitis C virus E2 antigen. Thus, CCR5 is likely to exert immunoregulatory effects in hepatitis C virus infection by preferentially recruiting CD8(+) T cells bearing the inhibitory NKG2A receptor to the liver.