MSc (Microbiology) / Department of Microbiology

According to WHO, diarrhea disease is the second leading cause of death worldwide for children under five. Around 525 000 children under five are killed every year by diarrhea. Most deaths from diarrhea occur among children less than 2 years of age living in South Asia and sub-Saharan Africa. With a comprehensive strategy that ensures all children in need receive critical prevention and treatment measures it is possible to save the lives of millions of children at risk of death from diarrhea. The aim of this study was to identify what kind of treatment has been given to under-five children with symptoms of diarrhea and what kind of care the respondent sought in Zambia, a low-income country. Method: A cross-sectional study with secondary data from Zambia Demographic and Health Survey (ZDHS). The first-born child under five with diarrhea the last two weeks was selected in this study, to analyze what kind of treatment was given when showing symptoms of diarrhea. Out of the first-born children (n=7048) we found that 1 289 children had diarrhea during the last two weeks before the survey. Result: Of children having diarrhea 80% received some kind of treatment. Thirty eight percent of the children with diarrhea symptoms was given zinc and 67% was given pre-packaged Oral Rehydration Solution (ORS). The first place to seek treatment for the child’s diarrhea was at the Government Health Center (66%)followed by 18% that sought treatment at the Government Health Post. Conclusion: This study showed that the treatment and care seeking behavior for caregivers to under-5 children with diarrhea is of public health concern. Less than half of the children receives zinc as a treatment and 67% receives prepacked ORS. There is a need for education and awareness on the efficacy of ORS and especially zinc in preventing diarrhoea mortality and contribute to the UN Sustainable Development Goals target 3.2. / <p>Betyg i Ladok 210314.</p>

Thesis (MCur)--Stellenbosch University, 2015. / ENGLISH ABSTRACT: Annually more than 3.5 million children worldwide, less than five years of age die of diarrhoea and acute lower respiratory-tract infections. Over the last two decades, the care of pre-school children outside of their homes has become more common in most parts of the world and has contributed to an increased risk of respiratory-tract and gastrointestinal infections in children. Children attending day-care centres are at an increased risk for diarrhoea and respiratory-tract infections and hands are the primary vehicle for transmission of infectious diseases. Thus, hand hygiene is essential for preventing and controlling of infection in the health care and community settings. Waterless hand sanitizer as an alternative to hand washing was investigated. Rinse-free hand sanitizer programmes in the community may be effective, safe and feasible. The aim of the study was to systematically appraise evidence on the effect of alcohol-based hand rub in the prevention of diarrhoea and acute respiratory-tract infection among children aged five years and below in community settings. The primary outcome of the study was to assess the incidence of respiratory-tract infection and diarrhea. Diarrhoea is defined by the World Health Organization as the excretion of three or more loose or liquid stools per day (or more frequent excretion than is normal for the individual). The secondary outcome was to assess mortality, admission to hospital and duration of hospital stay. A comprehensive search for relevant studies was conducted on the following databases from 1990 to 2014: EMBASE, MEDLINE, CINAHL, Google Scholar and Cochrane Central Register of Controlled Trials (CENTRAL). We searched the reference lists of all relevant articles and textbooks for more studies. Unpublished data previously presented at international and scientific meetings have been included in the review. Proceedings of international conferences on diarrhoea and respiratory-tract infection among children were searched for relevant articles. Subject experts were contacted. Two reviewers, Joelynn Steyn (JS) and Oswell Khondowe (OK) selected studies following a two-step study selection process. This review considered all published randomized controlled trials and quasi-experimental designs published from January 1990 to July 2014. The first step was the selection of studies based on titles and abstracts. Both reviewers selected eligible studies which met the set criteria. During the second step, both reviewers retrieved the full-text articles of the studies and assessed the methodological quality of the studies. Four studies were included in this review. The included studies met most of the quality assessment criteria as stipulated in the Cochrane risk assessment tool. Two studies were clusterrandomized controlled trials, one was a block randomized controlled trial and one was a randomized controlled trial. Disagreements were resolved by discussion and where a lack of consensus existed, consultation with a third reviewer occurred. The use of alcohol hand rub as compared to control interventions significantly reduced the incidence of diarrhoea in children (RR 0.79, 95% CI 0.63 to 0.99). Statistical heterogeneity was observed among the included studies (I²=69, p=0.04). However this review found no significant difference in respiratory-tract infections between intervention groups versus control as observed from the confidence interval (RR 0.98, 95% CI 0.90 to 1.07, p=0.63). The results should be interpreted with caution due to the limited number of studies conducted in communities with alcoholbased hand rub used by caregivers. Due to limited studies in this review, it makes it difficult to make strong conclusions on findings and to provide sufficient evidence to guide future research. We therefore recommend that more studies with high quality methodologies, using randomized controlled trial designs be conducted especially in poor resourced communities. / AFRIKAANSE OPSOMMING: Meer as 3.5 miljoen kinders jonger as vyf jaar oud sterf jaarliks wêreldwyd as gevolg van diarree en akute laer respiratoriese lugweginfeksies. Oor die laaste dekades het die versorging van voorskoolse kinders buite hul tuiste meer algemeen geword in die meeste dele van die wêreld wat bygedra het tot ’n risiko in die toename van respiratoriese en spysverteringskanaalinfeksies by kinders. Kinders by dagsorgsentrums het ’n groter risiko vir diarree en respiratoriese lugweë infeksies want die oordra van siekte-infeksies word veral deur die hande wat as die primêre bron daarvan beskou word, gesien. Dus is hand-higiëne noodsaaklik om infeksies in gesondheidsorg en gemeenskapsentrums te voorkom en te beheer. ’n Waterlose hande-ontsmettingsmiddel as ’n alternatief om hande te was, is ondersoek. Spoelvrye hande-ontsmettingsmiddel programme in die gemeenskap mag effektief, veilig en uitvoerbaar wees. Die doel van die studie was om sistematies die geslaagdheid van bewyse op die effek van alkoholgebaseerde hande-smeermiddel in die voorkoming van diarree en akute lugweginfeksies by kinders 5 jaar en jonger in gemeenskapsentrums te ondersoek. Die primêre uitkoms van die studie was om die voorkoms van respiratoriese lugweginfeksie en diarree te asseseer. Diarree soos gedefinieer deur die WGO is die uitskeiding van drie of meer los- of waterige stoelgange per dag (of meer gereelde uitskeiding wat normaal vir die individu is). Die sekondêre uitkoms was om mortaliteit, toelating tot die hospitaal en duur van hospitaalverblyf te assesseer. ’n Omvattende soektog vir relevante studies was op die volgende databasisse van 2004-2014 uitgevoer: EMBASE, MEDLINE, CINAHL, Google Scholar en Cochrane Sentrale Register van Beheerde Toetse. Ons het die verwysingslyste van alle relevante artikels en handboeke vir meer navorsingstudies nagegaan. Ongepubliseerde data wat voorheen aangebied is by internasionale en wetenskaplike vergaderings, is ingesluit in die oorsig. Bevindings by internasionale konferensies oor diarree en lugweginfeksies by kinders was nagegaan in die soektog na relevante artikels. Onderwerpdeskundiges was gekontak. Die twee navorsers, Joelynn Steyn (JS) en Oswell Khondowe (OK) het studies geselekteer deur ’n twee-stap studieselekteringsproses te volg. Die oorsig het alle gepubliseerde, ewekansige gekontroleerde proewe en kwasi-eksperimenteerde studies oorweeg tussen Januarie 1990 en Julie 2014. Die eerste stap was die selektering van studies gebaseer op hul titels en opsommings. Beide navorsers het geskikte studies slegs geselekteer as die studie aan die bepaalde kriteria voldoen het. Tydens die tweede stap het beide navorsers die volledige artikels geneem van die studies geselekteer en die gehalte van die metodologie geassesseer. Vierstudies is in die oorsig ingesluit. Die ingeslote studies het aan die meeste vereistes soos deur die Cochrane risiko assesseringsinstrument gestipuleer voldoen. Twee studies was groep-ewekansige gekontroleerde proewe, een was ’n blok ewekansige gekontroleerde proef en een was ’n ewekansige gekontroleerde proef. Verskille is opgelos deur bespreking en waar daar onderbreking in konsensus was, het ’n konsultasie met ’n derde beoordelaar plaasgevind. Die gebruik van ’n alkohol handreiniger, soos vergelyk met kontrole-intervensies, het die voorkoms van diarree in kinders beduidend verminder (RR 0.79, 95% CI 0.63 to 0.99). Statistiese heterogeniteit is egter tussen die ingeslote studies waargeneem (I²=69, p=0.04). Hierdie studie het geen beduidende verskille in lugweg infeksies tussen die intervensiegroepe teenoor die kontrolegroep gevind nie soos waargeneem uit die vertroue interval (RR0.98, 95% CI 0.90 to 1.07, p=0.63). Die resultate moet met omsigtigheid geïnterpreteer word as gevolg van die beperkte aantal studies wat in gemeenskappe uitgevoer was met alkohol-gebasseerde handreiniger wat deur versorgers gebruik word. As gevolg van die beperkende aantal studies in hierdie studie, is dit moeilik om sterk gevolgtrekkings te maak op bevindinge en om voldoende bewyse te gee om toekomstige navorsing te rig. Ons beveel dus aan dat meer studies met hoë kwaliteit metodologie en wat ewekansige gekontroleerde proef-ontwerpe gebruik, uitgevoer word, veral in swak-toegeruste gemeenskappe. (RR0.98, 95% CI 0.90 to 1.07, p=0.63). Die resultate moet met omsigtigheid geïnterpreteer word as gevolg van die beperkte aantal studies wat gedoen is in gemeenskappe met alkohol-

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INTRODUCCIÓN. Los leucocitos fecales son utilizados para identificar diarrea invasiva y decidir el uso de antibióticos. Se conoce poco sobre su utilidad en hospitales de países en desarrollo con procesos de laboratorio eficientes. Buscamos evaluar el rendimiento diagnóstico de la prueba en menores de 5 años con diarrea aguda. MATERIAL Y MÉTODOS. Estudio retrospectivo de registros clínicos y de laboratorio en el Hospital de Emergencias Pediátricas, Lima, Perú. Se evaluó los casos a los que se había solicitado sistemática e independientemente leucocitos fecales y coprocultivo. Se calculó sensibilidad, especificidad, valores predictivos, cocientes de probabilidad (CP) y la curva de características operativas del receptor (ROC). RESULTADOS. De 1,804 muestras fecales, 901 (49,9%) fueron positivos para uno o más enteropatógenos bacterianos. La sensibilidad (S), especificidad (E), y el CP positivo variaron para los diferentes umbrales: más de 5 leucocitos por campo (S: 93.2%, E: 21.9%, CP: 1.9), más de 20 (S: 88.4%, E: 34.8%, CP: 1.35), más de 50 (S: 74.9%, E: 56.7%, CP: 1.73), y más de 100 (S: 60.7%, E: 71.9%, CP: 2.17). El área bajo la curva ROC fue 0.69 (IC 95%: 0.67-0.72). CONCLUSIONES. El rendimiento de la prueba es sub-óptimo y continuar su uso rutinario en la práctica clínica no parece justificado, pues promueve el abuso de antibióticos y por otro lado aumenta el riesgo de pasar por alto pacientes con diarrea invasiva. Se necesita estudiar el rendimiento diagnóstico de datos epidemiológicos y clínicos combinados con leucocitos fecales o lactoferrina fecal, para identificar una aproximación más eficiente. / INTRODUCTION. Fecal leukocytes are widely used to identify invasive diarrhea and to make then the decision of prescribing or not antibiotics. This test has been hardly assessed in small hospitals of developing countries with efficient laboratory processes. We aimed to assess the diagnostic performance of different thresholds of fecal leukocytes in children under-five with acute diarrhea. MATERIAL AND METHODS. Retrospective study of clinical and laboratory records in the Pediatric Emergency Hospital, Lima, Peru. All cases with a stool culture and fecal leukocytes independently and systematically performed were studied. Sensitivity, specificity, predictive values, likelihood ratios (LR), and receiver operating characteristics (ROC) curves were calculated. RESULTS. Out of 1,804 stool samples assessed, 901 (49,9%) were positive for one or more bacterial entheropathogens. Sensitivity (Sn), specificity (Sp), and positive LR varied for different thresholds: more than 5 (S: 93.2%, Sp: 21.9%, LR+:), more than 20 (Sn: %, Sp: %, +LR: ), more than 50 (Sn: 74.9%, Sp: 56.7%, +LR: 1.73), and more than 100 fecal leukocytes per high power field (Sn: 60.7%, Sp: 71.9%, LR+: 2.17). The general area under the ROC curve was 0.69 (CI 95%: 0.67-0.72).

theresa.j.ochoa@uth.tmc.edu / Article / BACKGROUND: Enteropathogenic Escherichia coli (EPEC) strains are pediatric pathogens commonly isolated from both healthy and sick children with diarrhea in areas of endemicity. The aim of this study was to compare the bacterial load of EPEC isolated from stool samples from children with and without diarrhea to determine whether bacterial load might be a useful tool for further study of this phenomenon. METHODS: EPEC was detected by polymerase chain reaction (PCR) of colonies isolated on MacConkey plates from 53 diarrheal and 90 healthy children aged <2 years. DNA was isolated from stool samples by cetyltrimethylammonium bromide extraction. To standardize quantification by quantitative real-time PCR (qRT-PCR), the correlation between fluorescence threshold cycle and copy number of the intimin gene of EPEC E2348/69 was determined. RESULTS: The detection limit of qRT-PCR was 5 bacteria/mg stool. The geometric mean load in diarrhea was 299 bacteria/mg (95% confidence interval [CI], 77-1164 bacteria/mg), compared with 29 bacteria/mg (95% CI, 10-87 bacteria/mg) in control subjects (P = .016). Bacterial load was significantly higher in children with diarrhea than in control subjects among children <12 months of age (178 vs 5 bacteria/mg; P = .006) and among children with EPEC as the sole pathogen (463 vs 24 bacteria/mg; P = .006). CONCLUSIONS: EPEC load measured by qRT-PCR is higher in diarrheal than in healthy children. qRT-PCR may be useful to study the relationship between disease and colonization in settings of endemicity.

Introduction: Recently, Clostridium difficile has garnered both national and international attention, occurring with greater frequency, severity and resistance to standard treatments. In 2001, Clostridium difficile-associated disease, CDAD, became epidemic and its rate doubled from 2000 to 2003. As a substantial cause of healthcare-associated morbidity, CDAD is no longer just a pharmacotherapy nuisance. Specifically, the objectives of this research are to determine the incidence of both primary and secondary cases of CDAD among patients in the UHC database, determine how many primary cases had previous admissions with secondary cases of CDAD, and determine how many primary cases go on to be readmitted as primary or secondary cases.Methods: This cross-sectional study used the dataset from the University HealthSystem Consortium (UHC) covering the first quarter of 2002 through the second quarter of 2006, to identify the incidence of primary and secondary CDAD within hospitals participating in the UHC's Clinical Resource Manager (CRM) program. The UHC-CRM database was used to provide epidemiologic data in regards to CDAD cases, hospital-wide variables and patient characteristics. The data was analyzed using SAS statistical software.Results: During the time period studied, a total of 17,636 CDAD cases were identified out of 2,805,901 total hospital discharges. Of those, 80.4% or 14,178 fit the definition of primary with 3,454 cases labeled as secondary. The mean discharge rate of primary CDAD cases, per 1,000 adult discharges among CRM-participating hospitals per quarter, ranged from 3.14 (SD 2.33) to 6.72 (SD 4.0). The mean incidence of secondary CDAD per 1,000 adult discharges among CRM-participating hospitals per quarter, ranged from 0.79 (SD 0.74) to 1.59 (SD 0.86). The mean discharge rate of all CDAD cases, per 1,000 (adult discharges) among CRM-participating hospitals per quarter, ranged from 3.93 (SD 3.04) to 8.11 (SD 4.52). The rate of CDAD case discharges shows an increasing trend, more than doubling from 2002 to 2006. Among previous primary CDAD admissions, 7.75% were readmitted as a primary CDAD cases (p-value Conclusions: CDAD cases are on the rise, with all CDAD cases diagnosed on discharge from doubling from 2002 to 2006. There is no current national surveillance system for tracking CDAD cases. This study begins to shed light on the increasing incidence of CDAD cases within the hospital setting. By providing a baseline, future research can use this information to make more extensive tracking and surveillance systems for CDAD cases.

Master of Science / Department of Grain Science and Industry / Cassandra K. Jones / Animal food manufacturing facilities need to evaluate biological hazards within their facility due to their severity and probability to cause illness or injury in humans or animals. Control of biological hazards, including Salmonella and Porcine Epidemic Diarrhea Virus (PEDV), in animal food manufacturing facilities may require a preventative control to mitigate the risk of the hazard. Thermal processing is an effective point-in-time control, but does not prevent cross-contamination during drying, cooling, and packaging/load-out of animal food. Therefore, it may be appropriate to sanitize surfaces to prevent cross-contamination of animal food during manufacturing. The objective of the first experiment was to evaluate surface decontamination strategies for Porcine Epidemic Diarrhea Virus (PEDV) using chemical disinfectants to reduce viral RNA on various manufacturing surfaces. Concentrated liquid formaldehyde and sodium hypochlorite reduced the quantity of viral PEDV RNA on all tested surfaces. Rubber belting from a bucket elevator retained the most PEDV RNA, while the polyethylene tote bag retained the least. In the second experiment, surface decontamination was evaluated for Salmonella Typhimurium using liquid and dry chemical sanitizers on various manufacturing surfaces. Surfaces treated with concentrated commercial formaldehyde had no detectable Salmonella after treatment, and surfaces treated with medium chain fatty acids (MCFA) had at least a 4-log reduction compared to the control. The dry commercial acidulant, sodium bisulfate, was the most effective dry sanitizer tested, but had limited efficacy depending on surface type. Experiment 3 further tested the application of two chemical sanitizers against Salmonella Enteritidis on residual surface and feed contamination in pilot-scale mixers. Manufacturing sequence, but not treatment impacted feed and surface contamination of Salmonella Enteritidis. Specifically, there was Salmonella-positive residue in the batch of feed manufactured immediately after the positive control batch. However, no Salmonella residue was detected in batches of feed treated with either concentrated commercial essential oil blend or rice hulls treated with 10% MCFA. Low levels of Salmonella residues were observed from feed and surfaces manufactured after Sequence 1, but no residues were observed by Sequence 2. This data suggests that sequencing of feed during manufacturing can reduce Salmonella-positive contamination within animal food and on manufacturing surfaces, particularly after the second batch or with the use of chemical treatments. In summary, liquid sanitizers have been shown to be effective at reducing Salmonella spp. and PEDV contamination on a variety of animal food manufacturing surfaces, but application and practicality may be limited.

Orientador: Katia Denise Saraiva Bresciani / Banca: Wagner Luiz Ferreira / Banca: Jancarlo Ferreira Gomes / Resumo: Parasitos gastrintestinais são comuns em animais de companhia. Atualmente, os cães e gatos de estimação apresentam um importante papel entre os humanos, sendo até considerados como membros da família. Por conta deste laço estreito com os humanos, surgem as preocupações dos veterinários em proporcionar uma melhor qualidade de vida para animais, e também em relação a algumas doenças são consideradas de caráter zoonótico. Alguns protozoários e helmintos (Giardia spp., Cryptosporidium spp., Isospora spp., Toxocara spp. e Ancylostoma spp.) são comumente diagnosticados, apesar das medidas terapêuticas e profiláticas existentes. Várias destas enfermidades apresentam sinais clínicos diversos, sendo a diarreia comumente encontrada. As técnicas coproparasitológicas tem como finalidade auxiliar o médico veterinário para um diagnóstico preciso e consequentemente o tratamento adequado ao animal. O objetivo do nosso trabalho foi investigar a ocorrência dos parasitos gastrintestinais em animais filhotes domiciliados. / Abstract: Gastrointestinal parasites are common in pets. Pets play an important role among humans and lately they have been considered members of families Due to the fact of the closely contact between humans and animals, veterinarians worries about zoonotic risk of some parasitic diseases and to promote better life quality for both pets and tutors. Some gastrointestinal parasites (Giardia spp., Cryptosporidium spp., Isospora spp., Toxocara spp. and Ancylostoma spp.) are commonly diagnosed causing diarrhea, despite existing therapeutic and prophylactic evidence. The coproparasitological techniques have been helping veterinarians for an accurate diagnosis and consequently the appropriate treatment to the animal. The objective of this study was to investigate the occurrence of gastrointestinal parasites in domiciled dogs. / Mestre

Escherichia coli enteroagregativa (EAEC) é caracterizada pela expressão do padrão de adesão agregativa (AA) em células HEp-2 e sua classificação em típica e atípica se baseia na presença ou ausência de aggR, respectivamente. O gene shf de EAEC codifica uma proteína com similaridade a IcaB, relacionada à formação de biofilme. Os objetivos deste estudo foram verificar a prevalência de shf em 113 amostras de EAEC isoladas no Brasil e investigar o envolvimento de Shf na formação de biofilme e no padrão AA de EAEC típica e atípica. O gene shf foi detectado em 25 EAEC típicas e em 16 atípicas. As amostras shf+ foram analisadas quanto ao sorogrupo, adesão em células HEp-2 e formação de biofilme. Nenhuma dessas características foi associada a shf e a formação de biofilme foi detectada em todas essas amostras. A mutagênese em shf resultou na diminuição da capacidade de formação de biofilme e na incapacidade de adesão. Foi demonstrado que o gene shf é prevalente e que Shf participa do estabelecimento do padrão AA e de formação de biofilme em EAEC típica e atípica. / Enteroaggregative Escherichia coli (EAEC) is characterized by the expression of the aggregative adherence (AA) pattern to HEp-2 cells. The classification of EAEC in typical and atypical is based on the presence or absence of aggR, respectively. The shf gene encodes a protein with similarity to IcaB, associated with biofilm formation. The objectives of this study were to verify the prevalence of shf in 113 EAEC strains isolated in Brazil and investigate the role of the Shf in the establishment of the AA pattern and biofilm of typical and atypical EAEC. The shf gene was detected in 25 typical and in 16 atypical EAEC. The shf+ strains were analyzed regarding serogroups, adherence to HEp-2 cells and the ability to form biofilm. None of these characteristics was associated with the presence of shf and all strains were able to produce biofilm. The mutagenesis in shf reduced the biofilm formation and abolished the adherence to HEp-2 cells. This study demonstrated that shf is prevalent and Shf is involved in the AA and biofilm formation of typical and atypical EAEC.