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Introduction Tibial shaft fractures are a common presenting injury among the pediatric population. Asthma is also a common diagnosis that is frequently encountered in this population and has a significant impact on healthcare utilization, quality of life, and mortality. Given the high prevalence of these conditions and risks of peri-anesthetic respiratory complications, the purpose of this study was to evaluate an association between asthma and the incidence of 30-day postoperative complications following the surgical management of tibial shaft fractures in the pediatric population. Methods The National Surgical Quality Improvement Program-Pediatric database was used to identify pediatric patients who underwent surgical treatment for tibial shaft fractures from 2013-2019. Patients were categorized into two groups: patients with a history of asthma and patients without a history of asthma. Differences in patient demographics, comorbidities, and postoperative complications were assessed using bivariate and multivariate analyses. Results Of the 2,649 patients who underwent surgical treatment for tibial shaft fractures, 115 (4.3%) had asthma. Compared to those without asthma, patients with a history of asthma were more likely to have other medical comorbidities. After controlling for the differences in baseline characteristics between the two groups, patients with asthma had an increased risk of prolonged hospital stay (OR 5.78; 95% CI 1.67 to 20.00; p=0.006). Conclusion Pediatric patients being surgically treated for tibial shaft fractures with asthma had an increased risk of prolonged hospital stay. It is important that proper preoperative workup, perioperative care, and understanding of the implications of asthma on postoperative recovery are appreciated to reduce prolonged hospitalization lengths and minimize hospital costs associated with tibial shaft fracture surgery.

Whipple’s disease (WD) is a rare infection due to Tropheryma whipplei. Following in-vitro cultivation of T. whipplei, an indirect-immunofluorescence serological assay (IFA) was developed. We tested the hypothesis that this assay could be used to either identify WD patients, or rule out WD, in patients in whom the diagnosis is being considered, based on the antibody titers of their IgM and IgG antibody responses. In this small study fourteen WD patients and 22 healthy volunteers’ sera were obtained from across Australia. All specimens were coded and de-identified before testing. A patient with an IgG antibody titer of ≤1:16 may have WD [sensitivity 57% (8/14) and specificity close to 100% (22/22)]. High IgM antibody titers (≥1:256) were more common in WD patients [sensitivity 50% (7/14) and specificity 86% (19/22)] than in controls. The area under Receiver-Operator-Characteristic curve for IgG in the IFA assay was 0.84 (95% CI 0.69-1.00). At an IgG antibody titer of ≤1:16 the Youden’s index was 0.57. WD patients’ under-produce IgG antibody to T.whipplei but are more likely to over-produce IgM antibodies. This screening IFA serological assay may be clinically useful in detecting those with a possible diagnosis of WD. Patients with an IgG antibody titer of ≤1:16 and an IgM antibody titer of ≥1:256 may have WD and should proceed to a tissue biopsy and PCR for confirmation. Further validation of this assay, by increasing the sample size, by testing it in patients with non-WD disease and trialing in other countries should be undertaken.

Q fever (caused by Coxiella burnetii) is thought to have an almost world-wide distribution, but few countries have conducted national serosurveys. We measured Q fever seroprevalence using residual sera from diagnostic laboratories across Australia. Individuals aged 1–79 years in 2012–2013 were sampled to be proportional to the population distribution by region, distance from metropolitan areas and gender. A 1/50 serum dilution was tested for the Phase II IgG antibody against C. burnetii by indirect immunofluorescence. We calculated crude seroprevalence estimates by age group and gender, as well as age standardised national and metropolitan/non-metropolitan seroprevalence estimates. Of 2785 sera, 99 tested positive. Age standardised seroprevalence was 5.6% (95% confidence interval (CI 4.5%–6.8%), and similar in metropolitan (5.5%; 95% CI 4.1%–6.9%) and non-metropolitan regions (6.0%; 95%CI 4.0%–8.0%). More males were seropositive (6.9%; 95% CI 5.2%–8.6%) than females (4.2%; 95% CI 2.9%–5.5%) with peak seroprevalence at 50–59 years (9.2%; 95% CI 5.2%–13.3%). Q fever seroprevalence for Australia was higher than expected (especially in metropolitan regions) and higher than estimates from the Netherlands (2.4%; pre-outbreak) and US (3.1%), but lower than for Northern Ireland (12.8%). Robust country-specific seroprevalence estimates, with detailed exposure data, are required to better understand who is at risk and the need for preventive measures.

Objective Although most patients recover from acute Q fever, around 20% develop Q fever fatigue syndrome (QFS), a debilitating fatigue syndrome that lasts at least 6 months. This study investigated transcriptional profiles of circulating monocytes and circulating cytokines as a subsequent mirror of myeloid cell function, 1 and 6 months after an acute Q fever infection. Methods Total RNA of circulating monocytes was collected from 11 acute Q fever patients and 15 healthy controls, matched for age (±5 years) and sex. Samples were collected at a median of 27 days (baseline, interquartile range, 15–35 days) after the infection and again 6 months thereafter. Transcriptome analysis was performed using RNA sequencing. Additionally, concentrations of circulating interleukin (IL)-10, IL-1β, IL-1Ra, and IL-6 were measured in serum. Results At baseline, acute Q fever patients clearly show a differential transcriptional program compared with healthy controls. This is still the case at follow-up, albeit to a lesser extent. At baseline, a significant difference in levels of circulating IL-10 (P = .0019), IL-1β (P = .0067), IL-1Ra (P = .0008), and IL-6 (P = .0003) was seen. At follow-up, this difference had decreased for IL-10 (P = .0136) and IL-1Ra (P = .0017) and had become nonsignificant for IL-1β (P = .1139) and IL-6 (P = .2792). Conclusions We show that an acute Q fever infection has a long-term effect on the transcriptional program of circulating monocytes and, therefore, likely their myeloid progenitor cells, as well as concentrations of circulating IL-10, IL-1β, IL-1Ra, and IL-6., This study shows that an acute Q fever infection has a long-lasting effect on the transcriptional program of circulating monocytes and, therefore, likely their myeloid progenitor cells as well as concentrations of circulating interleukin (IL)-10, IL-1β, IL-1Ra, and IL-6.

Objectives To estimate the prevalence of exposure to the causative agent of Q fever (Coxiella burnetii) and of current infections among blood donors in Australia. Design, setting Cross-sectional study in metropolitan Sydney and Brisbane, and in non-metropolitan regions with high Q fever notification rates (Hunter New England in New South Wales; Toowoomba in Queensland). Participants Blood donors attending Red Cross collection centres during October 2014 - June 2015 who provided sera and completed a questionnaire on Q fever vaccination status, diagnosis and knowledge, and exposure history. Main outcome measures Age- and sex-standardised seroprevalence of phase II IgG antibodies to C. burnetii (indicating past exposure) and independent risk factors for seropositivity; presence of C. burnetii DNA (indicating current infection and risk of transmission by blood transfusion). Results 2740 donors (94.5% response rate) completed the questionnaire and supplied sera for analysis. Crude antibody seroprevalence was 3.6%. Standardised seroprevalence was higher in non-metropolitan than metropolitan regions (NSW, 3.7% v 2.8%; Queensland, 4.9% v 1.6%; statistically significant only in Queensland). Independent predictors of antibody seropositivity were regular contact with sheep, cattle, or goats (adjusted odds ratio [aOR], 5.3; 95% CI, 2.1-14), abattoir work (aOR, 2.2; 95% CI, 1.2-3.9), and assisting at an animal birth (aOR, 2.1; 95% CI, 1.2-3.6). Having lived in a rural area but having only rare or no contact with sheep, cattle or goats was itself a significant risk factor (v never lived rurally: aOR, 2.5; 95% CI, 1.1-5.9). 40% of people in groups recommended for vaccination were aware of the vaccine; 10% of people in these groups had been vaccinated. C. burnetii DNA was not detected in 1681 non-metropolitan samples, suggesting that transmission by blood donation is unlikely. Conclusions Given their exposure to multiple risk factors, vaccination against Q fever should be considered for all rural residents.

Worldwide there are few isolate collections of the intracellular bacterium Coxiella burnetii, due to the difficulties associated with working with the organism and the scarcity of suitable samples from which to attempt isolation. Particularly lacking are isolates from acute Q fever patients. The aim of this study was to evaluate whether the serum samples taken from patients with confirmed acute Q fever during the early stage of their disease represented a potential source of viable C. burnetii. Isolation was attempted from 65 of these samples by inoculation of the serum into Vero cell culture and was successful in 36 cases (55%). This high success rate was likely due to extended incubation of up to twelve weeks of the inoculated cultures, allowing the growth of the organism to levels detectable by PCR. Retrospective analysis of the time the sera was stored prior to inoculation into culture demonstrated that C. burnetii remained viable for 224 days in samples stored refrigerated and 371 days in samples stored frozen at -20 °C. These results demonstrate that standard serum samples taken from acute Q fever patients are a valuable source of new isolates of C. burnetii, with no special handling of the specimens required to maintain the organism's viability.

There is limited evidence of rickettsial diseases in Bhutan. We explored the contribution of rickettsioses as a cause of undifferentiated febrile illness in patients presenting to 14 Bhutanese hospitals from October 2014 to June 2015. Obvious causes of fever were excluded clinically. Clinico-demographic information and acute blood samples were collected. Samples were tested by immunofluorescence assay (IFA) and qPCR against scrub typhus group (STG), spotted fever group (SFG) and typhus group (TG) rickettsiae, and Q fever (QF). Of the 1044 patients, 539 (51.6%) were female and the mean age was 31.5 years. At least 159 (15.2%) of the patients had evidence of a concurrent rickettsial infection. Of these, 70 (6.7%), 46 (4.4%), 4 (0.4%), and 29 (2.8%) were diagnosed as acute infections with STG, SFG, TG, and QF respectively. Ten (1.0%) patients were seropositive for both SFG and TG. Seven of the 70 STG patients were positive by qPCR. Eschar (p < 0.001), myalgia (p = 0.003), and lymphadenopathy (p = 0.049) were significantly associated with STG, but no specific symptoms were associated with the other infections. Disease incidences were not different between age groups, genders, occupations, and districts, except for students with significantly lower odds of infection with STG (OR = 0.43; 95% CI = 0.20, 0.93; p = 0.031). Rickettsioses were responsible for at least 15% of undifferentiated febrile illnesses in Bhutan, scrub typhus being the commonest. Health authorities should ensure that health services are equipped to manage these infections.

Background With few studies conducted to date, very little is known about the epidemiology of rickettsioses in Bhutan. Due to two previous outbreaks and increasing clinical cases, scrub typhus is better recognized than other rickettsial infections and Q fever. Methodology A descriptive cross-sectional serosurvey was conducted from January to March 2015 in eight districts of Bhutan. Participants were 864 healthy individuals from an urban (30%) and a rural (70%) sampling unit in each of the eight districts. Serum samples were tested by microimmunofluorescence assay for rickettsial antibodies at the Australian Rickettsial Reference Laboratory. Results Of the 864 participants, 345 (39.9%) were males and the mean age of participants was 41.1 (range 13–98) years. An overall seroprevalence of 49% against rickettsioses was detected. Seroprevalence was highest against scrub typhus group (STG) (22.6%) followed by spotted fever group (SFG) rickettsia (15.7%), Q fever (QF) (6.9%) and typhus group (TG) rickettsia (3.5%). Evidence of exposure to multiple agents was also noted; the commonest being dual exposure to STG and SFG at 5%. A person’s likelihood of exposure to STG and SFG rickettsia significantly increased with age and farmers were twice as likely to have evidence of STG exposure as other occupations. Trongsa district appeared to be a hotspot for STG exposure while Punakha district had the lowest STG exposure risk. Zhemgang had the lowest exposure risk to SFG rickettsia compared to other districts. People living at altitudes above 2000 meters were relatively protected from STG infections but this was not observed for SFG, TG or QF exposure. Conclusion This seroprevalence study highlights the endemicity of STG and SFG rickettsia in Bhutan. The high seroprevalence warrants appropriate public health interventions, such as diagnostic improvements and clinical treatment guidelines. Future studies should focus on vector profiles, geospatial, bio-social and environmental risk assessment and preventive and control strategies., Author summary Rickettsial infections including scrub typhus and Q fever are not widely recognised in Bhutan although the country is situated in an endemic Asian region. With two recorded outbreaks, scrub typhus is known to occur but other rickettsial infections and Q fever have not been recorded. In this first seroprevalence study of rickettsial infections, an overall seroprevalence of 49% was detected against rickettsioses amongst 864 participants. Seroprevalence was highest against scrub typhus group (STG) (22.6%) followed by spotted fever group (SFG) rickettsia (15.7%), Q fever (QF) (6.9%) and typhus group (TG) rickettsia (3.5%). Evidence of exposure to multiple agents were also noted; the commonest being dual exposure to STG and SFG at 5%. A person’s likelihood of exposure to STG and SFG significantly increased with age and farmers were twice as likely to have evidence of STG exposure as other occupations. Trongsa district in central Bhutan appeared to be a hotspot for STG exposure. People living at altitudes above 2000 meters were relatively protected from STG infections but this was not observed for SFG, TG and QF exposure. The findings from this study may direct future research on rickettsioses in Bhutan. These neglected tropical diseases warrant specific public health interventions in Bhutan.

A group of 14 persons who live in an area of Australia endemic for the Australian paralysis tick, Ixodes holocyclus, and who were involved in regularly collecting and handling these ticks, was examined for antibodies to tick-transmitted bacterial pathogens. Five (36%) had antibodies to Coxiella burnetii, the causative agent of Q fever and three (21%) had antibodies to spotted fever group (SFG) rickettsiae (Rickettsia spp). None had antibodies to Ehrlichia, Anaplasma, Orientia, or Borrelia (Lymedisease) suggesting that they had not been exposed to these bacteria. A total of 149 I. holocyclus ticks were examined for the citrate synthase (gltA) gene of the SFG rickettsiae and the com1 gene of C. burnetii; 23 (15.4%) ticks were positive for Rickettsia spp. and 8 (5.6%) positive for Coxiella spp. Sequencing of fragments of the gltA gene and the 17 kDa antigen gene from a selection of the ticks showed 99% and 100% homology, respectively, to Rickettsia australis, the bacterium causing Queenslandtick typhus. Thus, it appears that persons bitten by I. holocyclus in NE NSW, Australia have an approximate one in six risk of being infected with R. australis. Risks of Q fever were also high in this region but this may have been due to exposure by aerosol from the environment rather than by tick bite. A subset of 74 I. holocyclus ticks were further examined for DNA from Borrelia spp., Anaplasma spp. and Ehrlichia spp. but none was positive. Some of these recognised human bacterial pathogens associated with ticks may not be present in this Australian tick species from northeastern New South Wales.

In July 2006, an Australian tourist returning from Dubai, in the United Arab Emirates (UAE), developed acute scrub typhus. Her signs and symptoms included fever, myalgia, headache, rash, and eschar. Orientia tsutsugamushi serology demonstrated a 4-fold rise in antibody titers in paired serum collections (1:512 to 1:8,192), with the sera reacting strongest against the Gilliam strain antigen. An Orientia species was isolated by the in vitro culture of the patient's acute blood taken prior to antibiotic treatment. The gene sequencing of the 16S rRNA gene ( rrs ), partial 56-kDa gene, and the full open reading frame 47-kDa gene was performed, and comparisons of this new Orientia sp. isolate to previously characterized strains demonstrated significant sequence diversity. The closest homology to the rrs sequence of the new Orientia sp. isolate was with three strains of O. tsutsugamushi (Ikeda, Kato, and Karp), with a nucleotide sequence similarity of 98.5%. The closest homology to the 47-kDa gene sequence was with O. tsutsugamushi strain Gilliam, with a nucleotide similarity of 82.3%, while the closest homology to the 56-kDa gene sequence was with O. tsutsugamushi strain TA686, with a nucleotide similarity of 53.1%. The molecular divergence and geographically unique origin lead us to believe that this organism should be considered a novel species. Therefore, we have proposed the name “ Orientia chuto ,” and the prototype strain of this species is strain Dubai, named after the location in which the patient was infected.

Poultry movement is known to contribute to the dissemination of highly pathogenic avian influenza (HPAI) viruses. In Northern Vietnam, the illegal trade of poultry from China is a source of concern and is considered as responsible for the regular introduction of new H5N1 viruses. The general objective of this study was to get a better understanding of this illegal trade (organization, volume, actors involved and drivers) to propose adequate preventive and control options. The information was also used to qualitatively evaluate the risk of exposure of susceptible poultry to HPAI H5N1 virus introduced from China by illegally traded poultry. We found that the main products imported from China are spent hens, day-old chicks (DOCs) and ducklings; spent hens being introduced in very large number. The drivers of this trade are multiple: economic (especially for spent hens) but also technical (demand for improved genetic potential for DOC and ducklings). Furthermore, these introductions also meet a high consumer demand at certain periods of the year. We also found that spatial dispersion of a batch of poultry illegally introduced from China is extensive and rapid, making any prediction of possible new outbreaks very hazardous. Finally, a risk mitigation plan should include measures to tackle the drivers of this trade or to legally organize it, to limit the threat to the local poultry sector. It is also essential for traders to be progressively better organized and biosecure and for hygienic practices to be enforced, as our study confirmed that at-risk behaviours are still very common among this profession.

Although many studies have reported the indirect immunofluorescence assay (IFA) to be more sensitive in detection of antibodies to Coxiella burnetii than the complement fixation test (CFT), the diagnostic sensitivity (DSe) and diagnostic specificity (DSp) of the assay have not been previously established for use in ruminants. This study aimed to validate the IFA by describing the optimization, selection of cutoff titers, repeatability, and reliability as well as the DSe and DSp of the assay. Bayesian latent class analysis was used to estimate diagnostic specifications in comparison with the CFT and the enzyme-linked immunosorbent assay (ELISA). The optimal cutoff dilution for screening for IgG and IgM antibodies in goat serum using the IFA was estimated to be 1:160. The IFA had good repeatability (>96.9% for IgG, >78.0% for IgM), and there was almost perfect agreement (Cohen's kappa > 0.80 for IgG) between the readings reported by two technicians for samples tested for IgG antibodies. The IFA had a higher DSe (94.8%; 95% confidence interval [CI], 80.3, 99.6) for the detection of IgG antibodies against C. burnetii than the ELISA (70.1%; 95% CI, 52.7, 91.0) and the CFT (29.8%; 95% CI, 17.0, 44.8). All three tests were highly specific for goat IgG antibodies. The IFA also had a higher DSe (88.8%; 95% CI, 58.2, 99.5) for detection of IgM antibodies than the ELISA (71.7%; 95% CI, 46.3, 92.8). These results underscore the better suitability of the IFA than of the CFT and ELISA for detection of IgG and IgM antibodies in goat serum and possibly in serum from other ruminants.

We recently found that the herpes simplex virus-1 (HSV-1) latency-associated transcript (LAT) results in exhaustion of virus-specific CD8+ T cells in latently-infected trigeminal ganglia (TG). In this study we sought to determine if this impairment may involve LAT directly and/or indirectly interfering with DC maturation. We found that a small number of HSV-1 antigen-positive DCs are present in the TG of latently-infected CD11c/eYFP mice; however, this does not imply that these DCs are acutely or latently infected. Some CD8+ T cells are adjacent to DCs, suggesting possible interactions. It has previously been shown that wild-type HSV-1 interferes with DC maturation. Here we show for the first time that this is associated with LAT expression, since compared to LAT(−) virus: (1) LAT(+) virus interfered with expression of MHC class I and the co-stimulatory molecules CD80 and CD86 on the surface of DCs; (2) LAT(+) virus impaired DC production of the proinflammatory cytokines IL-6, IL-12, and TNF-α; and (3) DCs infected in vitro with LAT(+) virus had significantly reduced the ability to stimulate HSV-specific CD8+ T cells. While a similar number of DCs was found in LAT(+) and LAT(−) latently-infected TG of CD11c/eYFP transgenic mice, more HSV-1 Ag-positive DCs and more exhausted CD8 T cells were seen with LAT(+) virus. Consistent with these findings, HSV-specific cytotoxic CD8+ T cells in the TG of mice latently-infected with LAT(+) virus produced less IFN-γ and TNF-α than those from TG of LAT(−)-infected mice. Together, these results suggest a novel immune-evasion mechanism whereby the HSV-1 LAT increases the number of HSV-1 Ag-positive DCs in latently-infected TG, and interferes with DC phenotypic and functional maturation. The effect of LAT on TG-resident DCs may contribute to the reduced function of HSV-specific CD8+ T cells in the TG of mice latently infected with LAT(+) virus.

Summary Considering the limited success of the recent herpes clinical vaccine trial [1], new vaccine strategies are needed. Infections with herpes simplex virus type 1 and type 2 (HSV-1 & HSV-2) in the majority of men and women are usually asymptomatic and results in lifelong viral latency in neurons of sensory ganglia (SG). However, in a minority of men and women HSV spontaneous reactivation can cause recurrent disease (i.e., symptomatic individuals). Our recent findings show that T cells from symptomatic and asymptomatic men and women (i.e. those with and without recurrences, respectively) recognize different herpes epitopes. This finding breaks new ground and opens new doors to assess a new vaccine strategy: mucosal immunization with HSV-1 & HSV-2 epitopes that induce strong in vitro CD4 and CD8 T cell responses from PBMC derived from asymptomatic men and women (designated here as “asymptomatic” protective epitopes”) could boost local and systemic “natural” protective immunity, induced by wild-type infection. Here we highlight the rationale and the future of our emerging “asymptomatic” T cell epitope-based mucosal vaccine strategy to decrease recurrent herpetic disease.

Of the various rickettsial diseases, only scrub typhus has been well documented in Papua New Guinea. A review of the historical literature confirms this. A serological survey was conducted on 113 antenatal patients presenting to a district hospital in Kokopo, East New Britain. Results suggested that a spotted fever rickettsial infection is common in this area with a seroprevalence of about 17% in young women. There was no evidence of scrub typhus or murine (endemic) typhus in the population sampled. Clinical implications of these findings are discussed.

Protective “asymptomatic” human CD8+ T-cell epitopes from herpes simplex virus type 1 and type 2 (HSV1 & HSV-2) protein antigens remain largely undefined. Here, we have identified immunodominant protective "asymptomatic" human CD8+ T cell epitopes on the HSV-1 gB that are restricted to HLA-A*0201, an haplotype that covers over 50% of the human population. The 904-amino acid HSV-1 (strain 17) gB sequence was screened for potential CD8+ T cell epitopes. Ten epitopes were identified, synthesized and tested for affinity to HLA-A*0201 molecule. Two of these peptide epitopes, gB441-449 and gB561-569, exhibited high affinity for soluble HLA-A*0201 molecules. Peptide-specific CD8+ T cell proliferation (CFSE dilution) and CD107a/b cytotoxic degranulation were demonstrated against the same two epitopes in 30 sequentially studied HLA-A*0201-positive, HSV-1 seropositive asymptomatic individuals (i.e. during control of herpetic disease), but not in symptomatic individuals (i.e. with frequent herpetic disease). A heterologous lipopeptide/rAdv5 prime/boost vaccine expressing these two asymptomatic gB CD8+ epitopes induced a strong and long lasting CD8+ T-cell protective immunity against ocular and genital herpes in a susceptible HLA-A*0201 transgenic mice. Human protective asymptomatic CD8+ T-cell epitopes on HSV-1 proteins should continue to provide new insight into immune correlates of protection and aid in the development of a safe and efficient immunotherapeutic vaccine for herpes.