- Email: [email protected]
Laboratory diagnosis of Zika virus infection
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PATHOLOGY 2017 ABSTRACT SUPPLEMENT
as whole genome sequencing and its potential role in the characterisation of resistant strains will also be discussed. INFECTIVE ENDOCARDITIS IN THE 21ST CENTURY Eugene Athan1,2 1 Barwon Health, and 2Deakin University, Geelong, Vic, Australia Background: The epidemiology of infective endocarditis (IE) has changed dramatically in the last 50 years. Historically oral streptococci infected patients with underlying rheumatic heart disease resulting in sub-acute presentations. Discussion: The contemporary picture is increasing characterised by elderly comorbid patients with prosthetic valves and cardiac devices. The microbiology is characterised predominantly by Staphylococci including S. aureus, Enterococci and yeast. Health care associated infection accounts for up to half of all cases. Antimicrobial resistance poses a significant challenge. Modern approaches to diagnosis including the importance of blood cultures and transesophageal echocardiography will be discussed. The application of routine multidisciplinary management including infectious disease, cardiology and cardiac surgery will be discussed. The increasing role of surgery and the optimal timing to improve patient outcomes will be reviewed. ORAL MICROBIOME IN HEALTH AND DISEASE: LESSONS FROM HIV Mahmoud A. Ghannoum OHARA/ACTG Mycology Unit, Center for Medical Mycology, Department of Dermatology, Case Western Reserve University, Cleveland, OH, United States The oral cavity is home to microbial (bacterial, fungal and viral) communities with important implications for human health and disease. A healthy oral microbiome should be dominated by ‘core microbiome’. Disturbance in this core microbiome is likely to be linked to oral disease. Using HIV as a disease model, we characterised the core oral mycobiome (COM) and bacteriome (COB) in HIV-infected patients (HIV-IP) and matched uninfected controls (HC) [12 HIV-infected patients and 12 uninfected controls (matched for age, sex, and ethnicity)]. Our data showed that the number of bacterial and fungal genera ranged between 8–14 and 1–9 per individual patient, among uninfected and HIV-infected participants, respectively. The core oral bacteriome (COB) comprised 14 genera (of which 13 were common) in HIV-infected and uninfected individuals. The core oral mycobiome (COM) differed between HIV-infected and uninfected individuals, with Candida being the predominant fungus in both groups. Among Candida species, C. albicans was the most common (58% in uninfected and 83% in HIV-infected participants). Correlation analyses showed 15 and 12 bacteria-fungi pairs were correlated significantly in samples from uninfected and HIV-infected patients, respectively. Interestingly, increase in Candida colonisation was associated with a concomitant decrease in the abundance of Pichia (generally considered to be a harmless yeast and used in crop bi-control), suggesting antagonism. Further studies showed that Pichia cells exhibited direct anti-Candida activity mediated by proteinaceous secretory factors that inhibited Candida virulence factors (adhesion,
Pathology (2017), 49(S1)
germination, and biofilm formation). Our results identified HIVassociated changes in the oral mycobiome, and demonstrated that normal fungal community interacts with Candida. Characterisation of both mycobiome and bacteriome should be included in future investigations into the role of the human microbiome in health and disease. Our findings may lead to the discovery of a novel antifungal agent for the prevention and treatment of these diseases. NEW ASSAY FOR BACTERIAL LOAD AND SEPSIS MORTALITY (BLISS STUDY) Jonathan Iredell1,2, Shereen Mohsin1, Amith Shetty1,3, Andrew Ginn1,2 and on behalf of the BLISS investigators 1 Centre for Critical Infection, Westmead Institute & Marie Bashir Institute, Westmead Hospital, 2Institute for Clinical Pathology and Medical Research, Pathology NSW, Westmead Hospital, and 3Department of Emergency Medicine, Westmead Hospital, Westmead, NSW, Australia Sepsis is the biggest killer in emergency department presentations and kills more Australians than breast cancer, prostate cancer and HIV/AIDS combined. It long ago overtook stroke and heart attack as a cause for admission to hospital and continues an upward trajectory. New sepsis definitions have done nothing to improve the recognition of sepsis and risk of death at point of presentation and there is urgent need for improvements in rapid diagnostics. Procalcitonin and serum lactate measurement are practical point of care tests that help define inflammatory response and tissue injury, but a direct detection of bacteraemia is a potentially helpful adjunct. An Australia-wide study of resuscitation of septic shock (the ARISE study) allowed a nested sub-study of 108 patients to be recruited, in whom bacterial DNA load was determined using multiplexed consensus 16srDNA targets. Bacterial DNA above 100 cfu equivalents/mL of blood predicts death in hospital and is congruent with the APACHE 2 score. It is independent of all other variables in logistic regression analysis and a stronger risk for death than inotrope requirement. Bacterial load commonly remained positive for at least 3 days for Enterobacteriaceae. Twice as sensitive as blood culture, results are available in <4 hours. LABORATORY DIAGNOSIS OF ZIKA VIRUS INFECTION Linda Hueston Arbovirus & Emerging Diseases Unit, CIDMLS-ICPMR Pathology West, NSW, Australia In 1947 a new virus was isolated from a Rhesus macaque monkey in the Zika Forest near Entebbe in Africa. The isolation was serendipitous as the monkey was a sentinel animal in a Yellow Fever surveillance project. One human case was reported a year later in a child who had a febrile illness and made a full recovery. Serologic evidence suggested the virus had infected humans in various parts of Africa and Asia but no obvious overt cases of disease had been recorded. But in 2007 an outbreak of illness occurred on Yap Island in Micronesia – initially considered to be dengue it was later discovered to be due to Zika virus and this represented the first incursion into the Pacific. In 2013 it had spread to French Polynesia where a few cases of microcephaly and Guillain-
ABSTRACTS
Barre syndrome were noted. Over the next 3 years it spread across the Pacific causing large outbreaks. But in 2015 it reached the Americas and caused explosive outbreaks with large numbers of microcephaly being recorded. Although Zika has been recognised for 70 years diagnostic testing is not routinely available. Indeed investigation of Zika infection was either an entirely clinical approach or involved testing in one or two arbovirus reference laboratories in the world using haemagglutination inhibition or neutralisation testing. In 2013 our laboratory imported Zika virus to develop in-house testing ability for routine diagnosis for the Asia Pacific region. We began testing using neutralisation and then developed IgG and IgM IFA assays for first line screening. The combination of these two assays were used to diagnose outbreaks across the Western Pacific and to provide diagnostic services within Australia and Asia. However, laboratory diagnosis of flavivirus infections is not straightforward due to inherent cross reactivity among closely related viruses. This feature complicates laboratory diagnosis and given these results are often used to determine whether or not to terminate pregnancies it is important to understand the limitations of testing. In 2016 Euroimmun released an IgG and IgM ELISA for Zika virus antibody detection utilising a recombinant non-structural antigen. The use of non-structural antigens is believed to provide an exquisite specificity in arboviral tests, however our investigation indicates this is not the case. In this presentation we will examine the difficulties in making a laboratory diagnosis within a virus family often described as an antigenic hall of mirrors and review some cases highlighting some of the diagnostic dilemmas. ZIKA VIRUS UPDATE – PUBLIC HEALTH AND ENVIRONMENTAL ISSUES Vicky Sheppeard, Sean Tobin Health Protection NSW, Sydney, NSW, Australia From February to November 2016 Zika virus infection was considered a Public Health Emergency of International Concern (PHEIC) by the World Health Organization (WHO), prompting a coordinated international response to better understand the risks posed to human populations from this infection. While no longer a PHEIC, the WHO still considers that Zika virus infection is a ‘significant enduring public health challenge’ requiring a ‘sustained programme of work with dedicated resources to address the long-term nature of the disease and its associated consequences’. During the first 10 months of 2016, 55 Zika virus infections were notified in Australia. Infections were reported as acquired in central/south America (26), Pacific Islands (24) and south east Asia (5). The country where most Australians reported contracting Zika virus infection was Tonga (11) related to an outbreak there between February and April 2016. This presentation will cover the emergence of Zika virus infection, review the evidence on its clinical presentation and complications, and the guidance developed for Australian clinicians and public in collaboration between public health agencies, microbiologists and obstetric clinicians.
S53
Environmental aspects of Zika virus infection, such as competent mosquito vectors and their distribution in Australia will also be described, including exotic mosquito response arrangements. UPDATE ON RHEUMATIC FEVER – NEW INSIGHTS INTO THE PATHOGENESIS OF RHEUMATIC FEVER AND RHEUMATIC HEART DISEASE Natkunam Ketheesan1,2 1 School of Public Health, Medical and Veterinary Sciences, and 2 Australian Institute of Health Medical Research, James Cook University, Townsville, NSW, Australia Annually it is estimated that Streptococcus pyogenes (group A streptococcus; GAS) is responsible for over 700 million cases of pharyngitis and pyoderma. If untreated, pharyngitis and pyoderma can lead to acute rheumatic fever and rheumatic heart disease (ARF/RHD). Globally ARF/RHD affects over 33 million people. Therefore, an effective and safe vaccine against GAS infections will have significant health benefits. To develop GAS antigen based vaccines, a good understanding of the mechanisms leading to pathology is important. ARF/RHD is caused by immune responses predominantly triggered by GAS M protein, a protein which has structural homology with host tissue proteins. The post streptococcal sequela triggered by an autoimmune response involves both antibodies and T cells that cross-react with host tissue proteins such as cardiac myosin. Therefore, it is critical that any vaccine approach based on GAS antigens minimises the possibility of inducing autoreactive B and T cell responses. ARF/RHD is uniquely human and an animal model that encompasses the immunopathological and functional features of ARF/RHD is required to test vaccines. The rat autoimmune valvulitis (RAV) model has been used to investigate multiple mechanisms involved in initiating and potentiating cardiac damage including the potential of streptococci other than GAS contributing to the development of ARF/RHD. UPDATE ON RHEUMATIC FEVER IN NEW ZEALAND Arlo Upton Labtests Auckland and Northland Pathology Laboratory, New Zealand Despite being a relatively affluent nation, New Zealand (NZ) has persistently high rates of acute rheumatic fever (ARF) disproportionally affecting Maori and Pacific people. In 2011 the NZ government launched the Rheumatic Fever Prevention Programme (RFPP) with the stated aims of achieving equity of incidence and outcomes of ARF between Maori/Pacific and other NZ children, and to achieve a reduction in the national incidence of ARF by twothirds, from 4.0 per 100,000 people in 2011 to 1.4 per 100,000 people by 2017. The three interventions of the RFPP are: (1) improve access to timely diagnosis and treatment for group A streptococcal pharyngitis; (2) increase awareness of ARF; and (3) reduce household crowding and transmission of group A streptococcus. The presentation will cover the epidemiology of ARF in NZ, and discuss issues arising from the RFPP and outcomes.
PATHOLOGY 2017 ABSTRACT SUPPLEMENT
as whole genome sequencing and its potential role in the characterisation of resistant strains will also be discussed. INFECTIVE ENDOCARDITIS IN THE 21ST CENTURY Eugene Athan1,2 1 Barwon Health, and 2Deakin University, Geelong, Vic, Australia Background: The epidemiology of infective endocarditis (IE) has changed dramatically in the last 50 years. Historically oral streptococci infected patients with underlying rheumatic heart disease resulting in sub-acute presentations. Discussion: The contemporary picture is increasing characterised by elderly comorbid patients with prosthetic valves and cardiac devices. The microbiology is characterised predominantly by Staphylococci including S. aureus, Enterococci and yeast. Health care associated infection accounts for up to half of all cases. Antimicrobial resistance poses a significant challenge. Modern approaches to diagnosis including the importance of blood cultures and transesophageal echocardiography will be discussed. The application of routine multidisciplinary management including infectious disease, cardiology and cardiac surgery will be discussed. The increasing role of surgery and the optimal timing to improve patient outcomes will be reviewed. ORAL MICROBIOME IN HEALTH AND DISEASE: LESSONS FROM HIV Mahmoud A. Ghannoum OHARA/ACTG Mycology Unit, Center for Medical Mycology, Department of Dermatology, Case Western Reserve University, Cleveland, OH, United States The oral cavity is home to microbial (bacterial, fungal and viral) communities with important implications for human health and disease. A healthy oral microbiome should be dominated by ‘core microbiome’. Disturbance in this core microbiome is likely to be linked to oral disease. Using HIV as a disease model, we characterised the core oral mycobiome (COM) and bacteriome (COB) in HIV-infected patients (HIV-IP) and matched uninfected controls (HC) [12 HIV-infected patients and 12 uninfected controls (matched for age, sex, and ethnicity)]. Our data showed that the number of bacterial and fungal genera ranged between 8–14 and 1–9 per individual patient, among uninfected and HIV-infected participants, respectively. The core oral bacteriome (COB) comprised 14 genera (of which 13 were common) in HIV-infected and uninfected individuals. The core oral mycobiome (COM) differed between HIV-infected and uninfected individuals, with Candida being the predominant fungus in both groups. Among Candida species, C. albicans was the most common (58% in uninfected and 83% in HIV-infected participants). Correlation analyses showed 15 and 12 bacteria-fungi pairs were correlated significantly in samples from uninfected and HIV-infected patients, respectively. Interestingly, increase in Candida colonisation was associated with a concomitant decrease in the abundance of Pichia (generally considered to be a harmless yeast and used in crop bi-control), suggesting antagonism. Further studies showed that Pichia cells exhibited direct anti-Candida activity mediated by proteinaceous secretory factors that inhibited Candida virulence factors (adhesion,
Pathology (2017), 49(S1)
germination, and biofilm formation). Our results identified HIVassociated changes in the oral mycobiome, and demonstrated that normal fungal community interacts with Candida. Characterisation of both mycobiome and bacteriome should be included in future investigations into the role of the human microbiome in health and disease. Our findings may lead to the discovery of a novel antifungal agent for the prevention and treatment of these diseases. NEW ASSAY FOR BACTERIAL LOAD AND SEPSIS MORTALITY (BLISS STUDY) Jonathan Iredell1,2, Shereen Mohsin1, Amith Shetty1,3, Andrew Ginn1,2 and on behalf of the BLISS investigators 1 Centre for Critical Infection, Westmead Institute & Marie Bashir Institute, Westmead Hospital, 2Institute for Clinical Pathology and Medical Research, Pathology NSW, Westmead Hospital, and 3Department of Emergency Medicine, Westmead Hospital, Westmead, NSW, Australia Sepsis is the biggest killer in emergency department presentations and kills more Australians than breast cancer, prostate cancer and HIV/AIDS combined. It long ago overtook stroke and heart attack as a cause for admission to hospital and continues an upward trajectory. New sepsis definitions have done nothing to improve the recognition of sepsis and risk of death at point of presentation and there is urgent need for improvements in rapid diagnostics. Procalcitonin and serum lactate measurement are practical point of care tests that help define inflammatory response and tissue injury, but a direct detection of bacteraemia is a potentially helpful adjunct. An Australia-wide study of resuscitation of septic shock (the ARISE study) allowed a nested sub-study of 108 patients to be recruited, in whom bacterial DNA load was determined using multiplexed consensus 16srDNA targets. Bacterial DNA above 100 cfu equivalents/mL of blood predicts death in hospital and is congruent with the APACHE 2 score. It is independent of all other variables in logistic regression analysis and a stronger risk for death than inotrope requirement. Bacterial load commonly remained positive for at least 3 days for Enterobacteriaceae. Twice as sensitive as blood culture, results are available in <4 hours. LABORATORY DIAGNOSIS OF ZIKA VIRUS INFECTION Linda Hueston Arbovirus & Emerging Diseases Unit, CIDMLS-ICPMR Pathology West, NSW, Australia In 1947 a new virus was isolated from a Rhesus macaque monkey in the Zika Forest near Entebbe in Africa. The isolation was serendipitous as the monkey was a sentinel animal in a Yellow Fever surveillance project. One human case was reported a year later in a child who had a febrile illness and made a full recovery. Serologic evidence suggested the virus had infected humans in various parts of Africa and Asia but no obvious overt cases of disease had been recorded. But in 2007 an outbreak of illness occurred on Yap Island in Micronesia – initially considered to be dengue it was later discovered to be due to Zika virus and this represented the first incursion into the Pacific. In 2013 it had spread to French Polynesia where a few cases of microcephaly and Guillain-
ABSTRACTS
Barre syndrome were noted. Over the next 3 years it spread across the Pacific causing large outbreaks. But in 2015 it reached the Americas and caused explosive outbreaks with large numbers of microcephaly being recorded. Although Zika has been recognised for 70 years diagnostic testing is not routinely available. Indeed investigation of Zika infection was either an entirely clinical approach or involved testing in one or two arbovirus reference laboratories in the world using haemagglutination inhibition or neutralisation testing. In 2013 our laboratory imported Zika virus to develop in-house testing ability for routine diagnosis for the Asia Pacific region. We began testing using neutralisation and then developed IgG and IgM IFA assays for first line screening. The combination of these two assays were used to diagnose outbreaks across the Western Pacific and to provide diagnostic services within Australia and Asia. However, laboratory diagnosis of flavivirus infections is not straightforward due to inherent cross reactivity among closely related viruses. This feature complicates laboratory diagnosis and given these results are often used to determine whether or not to terminate pregnancies it is important to understand the limitations of testing. In 2016 Euroimmun released an IgG and IgM ELISA for Zika virus antibody detection utilising a recombinant non-structural antigen. The use of non-structural antigens is believed to provide an exquisite specificity in arboviral tests, however our investigation indicates this is not the case. In this presentation we will examine the difficulties in making a laboratory diagnosis within a virus family often described as an antigenic hall of mirrors and review some cases highlighting some of the diagnostic dilemmas. ZIKA VIRUS UPDATE – PUBLIC HEALTH AND ENVIRONMENTAL ISSUES Vicky Sheppeard, Sean Tobin Health Protection NSW, Sydney, NSW, Australia From February to November 2016 Zika virus infection was considered a Public Health Emergency of International Concern (PHEIC) by the World Health Organization (WHO), prompting a coordinated international response to better understand the risks posed to human populations from this infection. While no longer a PHEIC, the WHO still considers that Zika virus infection is a ‘significant enduring public health challenge’ requiring a ‘sustained programme of work with dedicated resources to address the long-term nature of the disease and its associated consequences’. During the first 10 months of 2016, 55 Zika virus infections were notified in Australia. Infections were reported as acquired in central/south America (26), Pacific Islands (24) and south east Asia (5). The country where most Australians reported contracting Zika virus infection was Tonga (11) related to an outbreak there between February and April 2016. This presentation will cover the emergence of Zika virus infection, review the evidence on its clinical presentation and complications, and the guidance developed for Australian clinicians and public in collaboration between public health agencies, microbiologists and obstetric clinicians.
S53
Environmental aspects of Zika virus infection, such as competent mosquito vectors and their distribution in Australia will also be described, including exotic mosquito response arrangements. UPDATE ON RHEUMATIC FEVER – NEW INSIGHTS INTO THE PATHOGENESIS OF RHEUMATIC FEVER AND RHEUMATIC HEART DISEASE Natkunam Ketheesan1,2 1 School of Public Health, Medical and Veterinary Sciences, and 2 Australian Institute of Health Medical Research, James Cook University, Townsville, NSW, Australia Annually it is estimated that Streptococcus pyogenes (group A streptococcus; GAS) is responsible for over 700 million cases of pharyngitis and pyoderma. If untreated, pharyngitis and pyoderma can lead to acute rheumatic fever and rheumatic heart disease (ARF/RHD). Globally ARF/RHD affects over 33 million people. Therefore, an effective and safe vaccine against GAS infections will have significant health benefits. To develop GAS antigen based vaccines, a good understanding of the mechanisms leading to pathology is important. ARF/RHD is caused by immune responses predominantly triggered by GAS M protein, a protein which has structural homology with host tissue proteins. The post streptococcal sequela triggered by an autoimmune response involves both antibodies and T cells that cross-react with host tissue proteins such as cardiac myosin. Therefore, it is critical that any vaccine approach based on GAS antigens minimises the possibility of inducing autoreactive B and T cell responses. ARF/RHD is uniquely human and an animal model that encompasses the immunopathological and functional features of ARF/RHD is required to test vaccines. The rat autoimmune valvulitis (RAV) model has been used to investigate multiple mechanisms involved in initiating and potentiating cardiac damage including the potential of streptococci other than GAS contributing to the development of ARF/RHD. UPDATE ON RHEUMATIC FEVER IN NEW ZEALAND Arlo Upton Labtests Auckland and Northland Pathology Laboratory, New Zealand Despite being a relatively affluent nation, New Zealand (NZ) has persistently high rates of acute rheumatic fever (ARF) disproportionally affecting Maori and Pacific people. In 2011 the NZ government launched the Rheumatic Fever Prevention Programme (RFPP) with the stated aims of achieving equity of incidence and outcomes of ARF between Maori/Pacific and other NZ children, and to achieve a reduction in the national incidence of ARF by twothirds, from 4.0 per 100,000 people in 2011 to 1.4 per 100,000 people by 2017. The three interventions of the RFPP are: (1) improve access to timely diagnosis and treatment for group A streptococcal pharyngitis; (2) increase awareness of ARF; and (3) reduce household crowding and transmission of group A streptococcus. The presentation will cover the epidemiology of ARF in NZ, and discuss issues arising from the RFPP and outcomes.