Classification of new morbillivirus and jeilongvirus sequences from bats sampled in Brazil and Malaysia.

As part of a broad One Health surveillance effort to detect novel viruses in wildlife and people, we report several paramyxovirus sequences sampled primarily from bats during 2013 and 2014 in Brazil and Malaysia, including seven from which we recovered full-length genomes. Of these, six represent the first full-length paramyxovirid genomes sequenced from the Americas, including two that are the first full-length bat morbillivirus genome sequences published to date. Our findings add to the vast number of viral sequences in public repositories, which have been increasing considerably in recent years due to the rising accessibility of metagenomics. Taxonomic classification of these sequences in the absence of phenotypic data has been a significant challenge, particularly in the subfamily Orthoparamyxovirinae, where the rate of discovery of novel sequences has been substantial. Using pairwise amino acid sequence classification (PAASC), we propose that five of these sequences belong to members of the genus Jeilongvirus and two belong to members of the genus Morbillivirus. We also highlight inconsistencies in the classification of Tupaia virus and Mòjiang virus using the same demarcation criteria and suggest reclassification of these viruses into new genera. Importantly, this study underscores the critical importance of sequence length in PAASC analysis as well as the importance of biological characteristics such as genome organization in the taxonomic classification of viral sequences.

URSID γ-HERPESVIRUS TYPE 1-RELATED VIRUS IN CAPTIVE BORNEAN SUN BEARS (HELARCTOS MALAYANUS EURYSPILUS) IN SABAH, MALAYSIA.

The Bornean sun bear (Helarctos malayanus euryspilus) is the smallest subspecies of sun bear. Their numbers are declining, and more research is needed to better understand their health and biology. Forty-four bears housed at the Bornean Sun Bear Conservation Centre (BSBCC) in Sabah, Malaysia, were screened for known and novel viruses in November 2018. Ursid γ-herpesvirus type 1 (UrHV-1) is a herpesvirus that has been detected from swab samples of clinically healthy sun bears and biopsy samples of oral squamous cell carcinoma in sun bears. We detected an UrHV-1-related virus from throat and rectal swabs by molecular viral screening in samples from 15.9% of the sun bears at BSBCC. None of the bears with the UrHV-1-related virus in this study had oral lesions. There is no known report of UrHV-1 detection in the wild sun bear population, and its association with oral squamous cell carcinoma is not fully understood. Finding an UrHV-1-related virus in a rehabilitation center is a concern because conditions in captivity may contribute to spreading this virus, and there is the potential of introducing it into wild populations when a bear is released. This study demonstrates an urgent need to carry out similar surveillance for sun bears in captivity as well as those in the wild, to better understand the impact of captivity on the prevalence and spread of UrHV-1-related viruses. Positive bears also should be monitored for oral lesions to better understand whether there is a causal relationship.

The first reported cases of elephant endotheliotropic herpesvirus infectious haemorrhagic disease in Malaysia: case report.

BACKGROUND: Elephant endotheliotropic herpesvirus haemorrhagic disease (EEHV HD) is the leading cause of death in captive Asian elephant calves in Asia, North America, and Europe with a mortality rate of ~ 65% in calves that are under human care. Although EEHV HD was first found in elephant camps, more recently it was identified in wild populations which poses a greater threat to the elephant population. Deaths due to EEHV HD have been seen in wild elephants, but the in-situ prevalence and mortality rate is unknown. We report the first EEHV HD cases in Malaysia from 3 wild born endangered Bornean elephant calves from Sabah with known typical clinical signs. CASE PRESENTATION: The first calf died within 24 h of the onset of clinical signs; the second calf died within 12 h of the onset of clinical signs. The third calf succumbed within 72 h. Necropsies revealed that all 3 calves had similar presentations of EEHV HD but in the third calf with less severity. We conducted conventional polymerase chain reaction (cPCR) assays and found EEHV DNA at all 7 loci in the 3 calves; it was identified as EEHV1A, the virus type that has been found in most other reported cases. CONCLUSION: Typical EEHV HD clinical signs and the molecular confirmation of EEHV by cPCR and sequencing point to EEHV as the cause of death. Further genetic investigation of the strain is in progress.

No Evidence of Coronaviruses or Other Potentially Zoonotic Viruses in Sunda pangolins (Manis javanica) Entering the Wildlife Trade via Malaysia.

The legal and illegal trade in wildlife for food, medicine and other products is a globally significant threat to biodiversity that is also responsible for the emergence of pathogens that threaten human and livestock health and our global economy. Trade in wildlife likely played a role in the origin of COVID-19, and viruses closely related to SARS-CoV-2 have been identified in bats and pangolins, both traded widely. To investigate the possible role of pangolins as a source of potential zoonoses, we collected throat and rectal swabs from 334 Sunda pangolins (Manis javanica) confiscated in Peninsular Malaysia and Sabah between August 2009 and March 2019. Total nucleic acid was extracted for viral molecular screening using conventional PCR protocols used to routinely identify known and novel viruses in extensive prior sampling (> 50,000 mammals). No sample yielded a positive PCR result for any of the targeted viral families-Coronaviridae, Filoviridae, Flaviviridae, Orthomyxoviridae and Paramyxoviridae. In the light of recent reports of coronaviruses including a SARS-CoV-2-related virus in Sunda pangolins in China, the lack of any coronavirus detection in our 'upstream' market chain samples suggests that these detections in 'downstream' animals more plausibly reflect exposure to infected humans, wildlife or other animals within the wildlife trade network. While confirmatory serologic studies are needed, it is likely that Sunda pangolins are incidental hosts of coronaviruses. Our findings further support the importance of ending the trade in wildlife globally.

Spotted Fever Rickettsiosis in a Wildlife Researcher in Sabah, Malaysia: A Case Study.

We present evidence for a case of spotted fever rickettsiosis with severe complications in a young adult male. Although spotted fever group rickettsiae (SFGR) have been reported as the most prevalent cause of rickettsiosis in rural areas of Sabah, Malaysia since the 1980s, this is the first detailed case report of suspected SFGR in the state. Current data on the prevalence, type, and thorough clinical reports on complications of SFGR and other rickettsioses in Sabah is lacking and required to raise the awareness of such diseases. There is a need to emphasize the screening of rickettsioses to medical personnel and to encourage the use of appropriate antibiotics as early treatment for nonspecific febrile illnesses in this region. Suspected rickettsioses need to be considered as one of the differential diagnoses for patients presenting with acute febrile illness for laboratory investigations, and early treatment instituted.

Macacine Herpesvirus 1 in Long-Tailed Macaques, Malaysia, 2009-2011.

Macacine herpesvirus 1 (MaHV1; B virus) naturally infects macaques (Macaca spp.) and can cause fatal encephalitis in humans. In Peninsular Malaysia, wild macaques are abundant, and translocation is used to mitigate human-macaque conflict. Most adult macaques are infected with MaHV1, although the risk for transmission to persons who handle them during capture and translocation is unknown. We investigated MaHV1 shedding among 392 long-tailed macaques (M. fascicularis) after capture and translocation by the Department of Wildlife and National Parks in Peninsular Malaysia, during 2009-2011. For detection of MaHV1 DNA, PCR was performed on urogenital and oropharyngeal swab samples. Overall, 39% of macaques were shedding MaHV1 DNA; rates of DNA detection did not differ between sample types. This study demonstrates that MaHV1 was shed by a substantial proportion of macaques after capture and transport and suggests that persons handling macaques under these circumstances might be at risk for exposure to MaHV1.

Experimentally reduced root-microbe interactions reveal limited plasticity in functional root traits in Acer and Quercus.

BACKGROUND AND AIMS: Interactions between roots and soil microbes are critical components of below-ground ecology. It is essential to quantify the magnitude of root trait variation both among and within species, including variation due to plasticity. In addition to contextualizing the magnitude of plasticity relative to differences between species, studies of plasticity can ascertain if plasticity is predictable and whether an environmental factor elicits changes in traits that are functionally advantageous. METHODS: To compare functional traits and trait plasticities in fine root tissues with natural and reduced levels of colonization by microbial symbionts, trimmed and surface-sterilized root segments of 2-year-old Acer rubrum and Quercus rubra seedlings were manipulated. Segments were then replanted into satellite pots filled with control or heat-treated soil, both originally derived from a natural forest. Mycorrhizal colonization was near zero in roots grown in heat-treated soil; roots grown in control soil matched the higher colonization levels observed in unmanipulated root samples collected from field locations. KEY RESULTS: Between-treatment comparisons revealed negligible plasticity for root diameter, branching intensity and nitrogen concentration across both species. Roots from treated soils had decreased tissue density (approx. 10-20 %) and increased specific root length (approx. 10-30 %). In contrast, species differences were significant and greater than treatment effects in traits other than tissue density. Interspecific trait differences were also significant in field samples, which generally resembled greenhouse samples. CONCLUSIONS: The combination of experimental and field approaches was useful for contextualizing trait plasticity in comparison with inter- and intra-specific trait variation. Findings that root traits are largely species dependent, with the exception of root tissue density, are discussed in the context of current literature on root trait variation, interactions with symbionts and recent progress in standardization of methods for quantifying root traits.

The NOSE study (nasal ointment for Staphylococcus aureus eradication): a randomized controlled trial of monthly mupirocin in HIV-infected individuals.

BACKGROUND: HIV-positive patients at HELP/PSI, Inc, an in-patient drug rehabilitation center, had a high baseline prevalence of Staphylococcus aureus colonization (49%) and incidence of infection (17%) in a previous year-long study. METHODS: A randomized, double-blinded, placebo-controlled study was conducted to determine whether repeated nasal application of mupirocin ointment would decrease the odds of S. aureus nasal colonization in 100 HELP/PSI patients over an 8-month period. A 5-day course of study drug was given monthly, and colonization was assessed at baseline and 1 month after each treatment. S. aureus infection was a secondary outcome. RESULTS: In repeated-measures analysis, mupirocin reduced the odds of monthly S. aureus nasal colonization by 83% compared with placebo [adjusted odds ratio (ORadj) = 0.17; P < 0.0001]. Subjects colonized at study entry had a 91% reduction in subsequent colonization (ORadj = 0.09; P < 0.0001). Mupirocin also suppressed S. aureus colonization in subjects not colonized at baseline (ORadj = 0.23; P = 0.006). There was no difference in infection rates between the mupirocin and placebo groups (hazard ratio = 0.49, P = 0.29). CONCLUSIONS: Monthly application of nasal mupirocin significantly decreased S. aureus colonization in HIV patients in residential drug rehabilitation. Monthly mupirocin application has a potential role in long-term care settings or in HIV-positive patients with high rates of S. aureus colonization and infection.

A postinfluenza model of Staphylococcus aureus pneumonia.

BACKGROUND: Postinfluenza Staphylococcus aureus pneumonias are increasingly recognized as a major form of life-threatening infections. METHODS: A mouse model of postinfluenza S. aureus pneumonia was developed. Mice were intranasally infected with bacteria alone or bacteria plus virus. Infection was assessed by mouse survival, lung histopathology, bacterial density in the lungs, and cellular response to infection. RESULTS: Mice infected with both influenza virus and S. aureus showed higher mortality, greater lung parenchymal damage, and greater bacterial density at metastatic tissue sites than mice infected with only S. aureus. At 4 h, more polymorphonuclear leukocytes and fewer CD11c(+) cells were found in lung samples from mice infected with virus and bacteria than in those from mice infected with bacteria. alpha-Hemolysin and protein A were maximally expressed 4 h after infection, and Panton-Valentine leukocidin was maximally expressed 72 h after infection, with higher levels of alpha-hemolysin expression in mice infected with bacteria alone. Interferon gamma expression was higher in tissue collected from mice infected with virus plus bacteria than in those from bacteria-infected mice. CONCLUSIONS: The results from this model demonstrate diverse effects caused by antecedent influenza virus infection, which have a profound influence on the morbidity and mortality associated with S. aureus pneumonia.

Staphylococcus aureus in the community: colonization versus infection.

BACKGROUND: Antibiotic-resistant Staphylococcus aureus infections have increased dramatically in the community, yet S. aureus nasal colonization has remained stable. The objectives of this study were to determine if S. aureus colonization is a useful proxy measure to study disease transmission and infection in community settings, and to identify potential community reservoirs. METHODOLOGY/PRINCIPAL FINDINGS: Randomly selected households in Northern Manhattan, completed a structured social network questionnaire and provided nasal swabs that were typed by pulsed field gel electrophoresis to identify S. aureus colonizing strains. The main outcome measures were: 1) colonization with S. aureus; and 2) recent serious skin infection. Risk factor analyses were conducted at both the individual and the household levels; logistic regression models identified independent risks for household colonization and infection. RESULTS: 321 surveyed households contained 914 members. The S. aureus prevalence was 25% and MRSA was 0.4%. More than 40% of households were colonized. Recent antibiotic use was the only significant correlate for household colonization (p = .002). Seventy-eight (24%) households reported serious skin infection. In contrast with colonization, five of the six risk factors that increased the risk of skin infection in the household at the univariate level remained independently significant in multivariable analysis: international travel, sports participation, surgery, antibiotic use and towel sharing. S. aureus colonization was not significantly associated with serious skin infection in any analysis. Among multiperson households with more than one person colonized, 50% carried the same strain. CONCLUSIONS/SIGNIFICANCE: The lack of association between S. aureus nasal colonization and serious skin infection underscores the need to explore alternative venues or body sites that may be crucial to transmission. Moreover, the magnitude of colonization and infection within the household suggests that households are an underappreciated and substantial community reservoir.

Staphylococcus aureus colonization and infection in New York State prisons.

Methicillin-resistant Staphylococcus aureus is increasingly responsible for staphylococcal outbreaks in prison. There is limited information on the source of the outbreak strains, risk factors for infection, and transmission of these strains within a prison. We conducted a survey to determine the prevalence of nasal colonization with S. aureus in 2 New York State prisons. S. aureus isolates from clinical cultures collected from all New York State prisons during a 6-month period were compared with the colonizing strains. Analyses were conducted to determine whether prison-level characteristics were associated with colonization or infection with S. aureus. The colonization rate was 25.5% (124/487); 10.5% of the isolates were methicillin resistant, all were staphylococcal chromosomal cassette (SCC)mec type IV, and 61.5% were Panton Valentine leukocidin (PVL) positive. Surprisingly, 21.6% of the methicillin-susceptible isolates were also PVL positive. Of the clinical isolates, 48.3% were methicillin resistant, with 93.1% of the latter being SCCmec type IV and 48.3% being PVL positive. The predominant clone was USA 300. Prison-level risk factors for infection included the proportion of inmates with drug offenses, the length of inmate stay, and the jail from which inmates originated. This study suggests that both new and long-term inmates act as sources of S. aureus strains, with the more virulent of the latter preferentially being selected as pathogens.

Community-associated methicillin-resistant Staphylococcus aureus prevalence: how common is it? A methodological comparison of prevalence ascertainment.

BACKGROUND: Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infections are becoming increasingly prevalent. There is geographic variation in their reported prevalence across the United States; however, studies reporting on CA-MRSA prevalence also demonstrate great variability in their case-finding methodology. We conducted a study to see how three different methods to ascertain CA-MRSA prevalence would lead to different estimates. METHODS: Different methods were used to identify cases of CA-MRSA colonization and/or infection in New York City. Method 1: retrospective review of clinical and surveillance cultures identified through a hospital computer database. Method 2: prospective collection of surveillance cultures in the same hospital's emergency department. Method 3: prospective collection of surveillance cultures in a community setting. RESULTS: Differing values for CA-MRSA prevalence resulted depending on the method and denominator used. All nares cultures as the denominator led to prevalence estimates of 0.3%-0.6%; all S. aureus as the denominator led to rates of 1.2%-5%; all MRSA as the denominator led to estimates of 5.5%-50%. CONCLUSIONS: A comparison of three methods revealed that variability in case-finding methodologies can lead to different prevalence estimates. Key factors to consider when comparing CA-MRSA rates include the definition of CA-MRSA, choice of denominator, and method and setting of sample collection.

SdrF, a Staphylococcus epidermidis surface protein, binds type I collagen.

Staphylococcus epidermidis is the leading cause of device-related infections. These infections require an initial colonization step in which S. epidermidis adheres to the implanted material. This process is usually mediated by specific bacterial surface proteins and host factors coating the foreign device. Some of these surface proteins belong to the serine-aspartate repeat (Sdr) family, which includes adhesins from Staphyloccus aureus and S. epidermidis. Using a heterologous expression system in Lactococcus lactis to overcome possible staphylococcal adherence redundancy we observed that one of these Sdr proteins, SdrF, mediates binding to type I collagen when present on the lactococcal cell surface. We used lactococcal recombinant strains, a protein-protein interaction assay and Western ligand blot analysis to demonstrate that this process occurs via the B domain of SdrF and both the alpha1 and alpha2 chains of type I collagen. It was also found that a single B domain repeat of S. epidermidis 9491 retains the capacity to bind to type I collagen. We demonstrated that the putative ligand binding N-terminal A domain does not bind to collagen which suggests that SdrF might be a multiligand adhesin. Antibodies directed against the B domain significantly reduce in vitro adherence of S. epidermidis to immobilized collagen.

Cellular coating of the left ventricular assist device textured polyurethane membrane reduces adhesion of Staphylococcus aureus.

OBJECTIVE: Infections are among the most common and serious complications of ventricular assist device implantation. These infections generally occur within the first 2 months after surgery. The basis for this high incidence of infection is not well established, so a murine intravascular infection model was developed with aortic implantation of the textured polyurethane patch material currently used in HeartMate ventricular assist devices (Thoratec Corporation Pleasanton, Calif). METHODS: Polyurethane patch material was placed in the wall of the mouse descending aorta. Mice were then infected with Staphylococcus aureus 1 or 14 days after implantation. In vitro adhesion studies were conducted with polyurethane membranes coated with endothelial cells and membranes coated with fibrinogen. RESULTS: Mice were susceptible to infection in both dose- and time-dependent fashions. The patch material was significantly more susceptible to infection at day 1 than day 14. Immunohistologic and morphologic studies demonstrated that the CD31+ cells deposited on the membrane surface phenotypically appeared to be endothelial cells. In vitro adhesion studies of polyurethane membranes coated with endothelial cells showed them to be less susceptible to S. aureus binding than were membranes coated with fibrinogen. CONCLUSION: Textured polyurethane membranes are less susceptible to infection as cellular deposition occurs. The time frame within which these membranes become populated with cellular material is consistent with the time-dependent clinical incidence of infection. Cellular coating of polyurethane may provide a strategy for reducing the risk of infection.

Staphylococcus aureus colonization and the risk of infection in critically ill patients.

OBJECTIVE: To evaluate the relationship between Staphylococcus aureus nasal and tracheal colonization and infection in medical intensive care unit (MICU) patients. The primary outcome was the incidence of S. aureus infection in colonized versus non-colonized patients. DESIGN: Prospective, observational cohort study. Patients admitted to the MICU during the study period were screened for S. aureus nasal and tracheal colonization by culture and a PCR assay twice weekly. Demographic, clinical, and microbiologic data were collected in the MICU and for 30 days after discharge. PFGE and antibiotic susceptibility testing were performed on all S. aureus nasal, tracheal, and clinical isolates. RESULTS: Twenty-three percent of patients (47 of 208) were nasally colonized with S. aureus. Twenty-four percent of these patients developed S. aureus infections versus 2% of non-colonized patients (P < .01). Nine of 11 patients with both nasal colonization and infection were infected by their colonizing strain. Two of 47 nasally colonized patients developed an infection with a different strain of S. aureus. Fifty-three percent of intubated patients with nasal colonization (10 of 19) had tracheal colonization with S. aureus as opposed to 4.9% of intubated, non-colonized patients (3 of 61) (P < .01). Parenteral antibiotics were ineffective at clearing nasal colonization. PCR detected S. aureus colonization (nasal and tracheal) within 6.5 hours with a sensitivity of 83% and a specificity of 99%. CONCLUSIONS: The incidence of S. aureus infection was significantly elevated in nasally colonized MICU patients. Techniques to rapidly detect colonization in this population may make targeted topical prevention strategies feasible.