Bacterial and fungal secondary infections occurring in COVID-19 patients followed in intensive care: a retrospective study.

INTRODUCTION: We aimed to investigate the effects of secondary bacterial and fungal infections on patient outcomes in patients followed up in the intensive care unit (ICU) due to coronavirus disease 2019 (COVID-19). METHODOLOGY: We retrospectively analyzed reverse transcriptase polymerase chain reaction (RT-PCR) positive COVID-19 patients followed in the ICU of our hospital between March 2020 and June 2021, using the hospital information system. Demographic data, pathogens causing a secondary infection, onset time of secondary infection, and patient outcomes were recorded. RESULTS: A total of 251 RT-PCR positive patients who met the inclusion criteria were evaluated. The mean length of stay (LOS) in the ICU was 13.3 ± 9.6 days. During this period, 165 (65.7%) patients died. When blood, urine, respiratory tract, and catheter cultures were examined, the number of patients with growth in at least one culture was 129 (51.4%). There was growth in a total of 227 cultures. The highest culture positivity rate was observed in respiratory tract samples (n = 94, 41.4%). Gram-negative bacterial pathogens (n = 130, 58.4%) predominated. Candida spp. was more frequent in urine cultures. The median day of the occurrence of secondary infection was 10 (range: 6-15). Patients who developed secondary infection had a longer LOS and higher mortality rate than patients who did not (p < 0.001). CONCLUSIONS: Gram-negative secondary infections, predominantly in respiratory tract cultures, occurred in COVID-19 patients followed in the ICU. As a result, the LOS was prolonged and mortality rates increased.

Adhesion of Klebsiella oxytoca to bladder or lung epithelial cells is promoted by the presence of other opportunistic pathogens.

The intestinal and respiratory tracts of healthy individuals serve as habitats for a diverse array of microorganisms, among which Klebsiella oxytoca holds significance as a causative agent in numerous community- and hospital-acquired infections, often manifesting in polymicrobial contexts. In specific circumstances, K. oxytoca, alongside other constituents of the gut microbiota, undergoes translocation to distinct physiological niches. In these new environments, it engages in close interactions with other microbial community members. As this interaction may progress to co-infection where the virulence of involved pathogens may be promoted and enhance disease severity, we investigated how K. oxytoca affects the adhesion of commonly co-isolated bacteria and vice versa during co-incubation of different biotic and abiotic surfaces. Co-incubation was beneficial for the adhesion of at least one of the two co-cultured strains. K. oxytoca enhanced the adhesion of other enterobacteria strains to polystyrene and adhered more efficiently to bladder or lung epithelial cell lines in the presence of most enterobacteria strains and S. aureus. This effect was accompanied by bacterial coaggregation mediated by carbohydrate-protein interactions occurring between bacteria. These interactions occur only in sessile, but not planktonic populations, and depend on the features of the surface. The data are of particular importance for the risk assessment of the urinary and respiratory tract infections caused by K. oxytoca, including those device-associated. In this paper, we present the first report on K. oxytoca ability to acquire increased adhesive capacities on epithelial cells through interactions with common causal agents of urinary and respiratory tract infections.

The characteristics of microbiome in the upper respiratory tract of COVID-19 patients.

BACKGROUND: Co-infection with other pathogens in coronavirus disease 2019 (COVID-19) patients exacerbates disease severity and impacts patient prognosis. Clarifying the exact pathogens co-infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is premise of the precise treatment for COVID-19 patients. METHODS: Sputum samples were collected from 17 patients in the COVID-19 positive group and 18 patients in the COVID-19 negative group. DNA extraction was performed to obtain the total DNA. Sequencing analysis using 16S and ITS rRNA gene was carried out to analyze the composition of bacterial and fungal communities. Meanwhile, all the samples were inoculated for culture. RESULTS: We did not observe significant differences in bacterial composition between the COVID-19 positive and negative groups. However, a significantly higher abundance of Candida albicans was observed in the upper respiratory tract samples from the COVID-19 positive group compared to the COVID-19 negative group. Moreover, the Candida albicans strains isolated from COVID-19 positive group exhibited impaired secretion of aspartyl proteinases. CONCLUSION: COVID-19 positive patients demonstrate a notable increase in the abundance of Candida albicans, along with a decrease in the levels of aspartyl proteinases, indicating the alteration of microbiota composition of upper respiratory tract.

Impact of respiratory viral infections on nasopharyngeal pneumococcal colonization dynamics in children.

PURPOSE OF REVIEW: Prevention of acute respiratory illnesses (ARI) in children is a global health priority, as these remain a leading cause of pediatric morbidity and mortality throughout the world. As new products and strategies to prevent respiratory infections caused by important pathogens such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza, respiratory syncytial virus and pneumococcus are advancing, increasing evidence suggests that these and other respiratory viruses and pneumococci may exhibit interactions that are associated with altered colonization and disease dynamics. We aim to review recent data evaluating interactions between respiratory viruses and pneumococci in the upper respiratory tract and their potential impact on pneumococcal colonization patterns and disease outcomes. RECENT FINDINGS: While interactions between influenza infection and subsequent increased susceptibility and transmissibility of colonizing pneumococci have been widely reported in the literature, emerging evidence suggests that human rhinovirus, SARS-CoV-2, and other viruses may also exhibit interactions with pneumococci and alter pneumococcal colonization patterns. Additionally, colonizing pneumococci may play a role in modifying outcomes associated with respiratory viral infections. Recent evidence suggests that vaccination with pneumococcal conjugate vaccines, and prevention of colonization with pneumococcal serotypes included in these vaccines, may be associated with reducing the risk of subsequent viral infection and the severity of the associated illnesses. SUMMARY: Understanding the direction and dynamics of viral-pneumococcal interactions may elucidate the potential effects of existing and emerging viral and bacterial vaccines and other preventive strategies on the health impact of these important respiratory pathogens.

Prevalence and risk factors associated with multidrug-resistant bacteria in COVID-19 patients.

Bacterial coinfection among patients with confirmed coronavirus disease 2019 (COVID-19) is a critical medical concern that increases the disease severity and mortality rate. The current study is aimed at evaluating the effects of bacterial coinfections among COVID-19 patients, especially in relation to degree of severity and mortality. A retrospective study was conducted for patients with positive COVID-19 test, admitted to a regional COVID-19 hospital in Jeddah, Saudi Arabia, between May and August 2020. A specimen (e.g., blood, urine, or sputum) was collected from patients with confirmed COVID-19, and was cultured to determine bacterial coinfection caused by multidrug resistant (MDR) bacteria. COVID-19 patients were categorized into 2 groups based on the result of bacterial coinfection culture, as COVID-19 patients with coinfection and COVID-19 patients without coinfection. Independent sample t test or Mann-Whitney U test was used to compare age and hospitalization period between these groups. In addition, binominal logistic regression was applied to identify risk factors associated with mortality and bacterial coinfection. The study included 342 patients with laboratory confirmed COVID-19. Eighty (23.3%) patients were diagnosed with bacterial coinfection, while the remaining 262 (76.6%) patients did not test positive for bacterial coinfection. Length of hospital stay was prolonged among COVID-19 patients diagnosed with bacterial coinfection (16.01 ±â€…11.36 days) when compared with patients without bacterial coinfection (6.5 ±â€…6.12 days). Likewise, the mortality rate was significantly higher among COVID-19 patients with bacterial coinfection (90%) compared to those without bacterial coinfection (49.2%). Gram-negative bacteria were predominant compared to gram-positive, as Klebsiella pneumoniae (35 [43.8%]) and Acinetobacter baumanni (32 [40%]). On the other hand, Staphylococcus aureus (4 [5%]), Enterococcus faecalis (1 [1.3%]), and Enterococcus faecium (1 [1.3%]) were identified as gram-positive bacterial species from recruited patients. The findings of the current study showed that prolong hospitalization is the main risk factor associated with bacterial coinfection and death. Thus, health care providers should minimize hospitalization as well as following a continuous monitoring for bacterial coinfection among COVID-19 patients, to control the spread of infection and reducing the severity and mortality rate among COVID-19 patients.

Risk factors, outcomes, and epidemiological and etiological study of hospitalized COVID-19 patients with bacterial co-infection and secondary infections.

BACKGROUND: As a common complication of viral respiratory tract infection, bacterial infection was associated with higher mortality and morbidity. Determining the prevalence, culprit pathogens, outcomes, and risk factors of co-infection and secondary infection occurring in hospitalized patients with coronavirus disease 2019 (COVID-19) will be beneficial for better antibiotic management. METHODS: In this retrospective cohort research, we assessed clinical characteristics, laboratory parameters, microbiologic results, and outcomes of laboratory-confirmed COVID-19 patients with bacterial co-infection and secondary infection in West China Hospital from 2022 December 2nd to 2023 March 15th. RESULTS: The incidence of bacterial co-infection and secondary infection, as defined by positive culture results of clinical specimens, was 16.3% (178/1091) and 10.1% (110/1091) respectively among 1091 patients. Acinetobacter, Klebsiella, and Pseudomonas were the most commonly identified bacteria in respiratory tract samples of COVID-19 patients. In-hospital mortality of COVID-19 patients with co-infection (17.4% vs 9.5%, p = 0.003) and secondary infection (28.2% vs 9.5%, p < 0.001) greatly exceeded that of COVID-19 patients without bacterial infection. Cardiovascular disease (1.847 (1.202-2.837), p = 0.005), severe COVID-19 (1.694 (1.033-2.778), p = 0.037), and critical COVID-19 (2.220 (1.196-4.121), p = 0.012) were proved to be risk factors for bacterial co-infection, while only critical COVID-19 (1.847 (1.202-2.837), p = 0.005) was closely related to secondary infection. CONCLUSIONS: Bacterial co-infection and secondary infection could aggravate the disease severity and worsen clinical outcomes of COVID-19 patients. Notably, only critical COVID-19 subtype was proved to be an independent risk factor for both co-infection and secondary infection. Therefore, standard empirical antibiotics was recommended for critically ill COVID-19 rather than all the inpatients according to our research.

Helicobacter pylori and Epstein-Barr virus infection in cell polarity alterations.

The asymmetrical distribution of the cellular organelles inside the cell is maintained by a group of cell polarity proteins. The maintenance of polarity is one of the vital host defense mechanisms against pathogens, and the loss of it contributes to infection facilitation and cancer progression. Studies have suggested that infection of viruses and bacteria alters cell polarity. Helicobacter pylori and Epstein-Barr virus are group I carcinogens involved in the progression of multiple clinical conditions besides gastric cancer (GC) and Burkitt's lymphoma, respectively. Moreover, the coinfection of both these pathogens contributes to a highly aggressive form of GC. H. pylori and EBV target the host cell polarity complexes for their pathogenesis. H. pylori-associated proteins like CagA, VacA OipA, and urease were shown to imbalance the cellular homeostasis by altering the cell polarity. Similarly, EBV-associated genes LMP1, LMP2A, LMP2B, EBNA3C, and EBNA1 also contribute to altered cell asymmetry. This review summarized all the possible mechanisms involved in cell polarity deformation in H. pylori and EBV-infected epithelial cells. We have also discussed deregulated molecular pathways like NF-κB, TGF-ß/SMAD, and ß-catenin in H. pylori, EBV, and their coinfection that further modulate PAR, SCRIB, or CRB polarity complexes in epithelial cells.

Porcine circovirus type 2 and Glaesserella parasuis serotype 4 co-infection activates Snail1 to disrupt the intercellular junctions and facilitate bacteria translocation across the tracheal epithelium.

Clinically, Porcine circovirus type 2 (PCV2) often causes disease through coinfection with other bacterial pathogens, including Glaesserella parasuis (G. parasuis), which causes high morbidity and mortality. However, the mechanism of PCV2 and G. parasuis serotype 4 (GPS4) co-infection is still not fully understood. In this study, swine tracheal epithelial cells (STEC) were used as a barrier model, and our results showed that PCV2 infection increased the adhesion of GPS4 to STEC, while decreasing the levels of ZO-1, Occludin and increasing tracheal epithelial permeability, and ultimately facilitated GPS4 translocation. Snail1 is a transcriptional repressor, and has been known to induce epithelial-to-mesenchymal transition (EMT) during development or in cancer metastasis. Importantly, we found that Snail1, as a transcriptional repressor, was crucial in destroying the tracheal epithelial barrier induced by PCV2, GPS4, PCV2 and GPS4 coinfection. For the first time, we found that PCV2, GPS4, PCV2 and GPS4 coinfection cross-activates TGF-ß and p38/MAPK signaling pathways to upregulate the expression of Snail1, down-regulate the levels of ZO-1 and Occludin, and thus disrupt the integrity of tracheal epithelial barrier then promoting GPS4 translocation. Finally, PCV2 and GPS4 co-infection also can activate TGF-ß and p38/MAPK signaling pathways in vivo and upregulate Snail1, ultimately down-regulating the expression of ZO-1 and Occludin. Our study elucidates how PCV2 infection promotes GPS4 to breach the tracheal epithelial barrier and aggravate clinical manifestations.

Solving polymicrobial puzzles: evolutionary dynamics and future directions.

Polymicrobial infections include various microorganisms, often necessitating different treatment methods than a monomicrobial infection. Scientists have been puzzled by the complex interactions within these communities for generations. The presence of specific microorganisms warrants a chronic infection and impacts crucial factors such as virulence and antibiotic susceptibility. Game theory is valuable for scenarios involving multiple decision-makers, but its relevance to polymicrobial infections is limited. Eco-evolutionary dynamics introduce causation for multiple proteomic interactions like metabolic syntropy and niche segregation. The review culminates both these giants to form evolutionary dynamics (ED). There is a significant amount of literature on inter-bacterial interactions that remain unsynchronised. Such raw data can only be moulded by analysing the ED involved. The review culminates the inter-bacterial interactions in multiple clinically relevant polymicrobial infections like chronic wounds, CAUTI, otitis media and dental carries. The data is further moulded with ED to analyse the niche colonisation of two notoriously competitive bacteria: S.aureus and P.aeruginosa. The review attempts to develop a future trajectory for polymicrobial research by following recent innovative strategies incorporating ED to curb polymicrobial infections.

Influenza enhances host susceptibility to non-pulmonary invasive Streptococcus pyogenes infections.

Streptococcus pyogenes (group A streptococcus; GAS) causes a variety of invasive diseases (iGAS) such as bacteremia, toxic shock syndrome, and pneumonia, which are associated with high mortality despite the susceptibility of the bacteria to penicillin ex vivo. Epidemiologic studies indicate that respiratory influenza virus infection is associated with an increase in the frequency of iGAS diseases, including those not directly involving the lung. We modified a murine model of influenza A (IAV)-GAS superinfection to determine if viral pneumonia increased the susceptibility of mice subsequently infected with GAS in the peritoneum. The results showed that respiratory IAV infection increased the morbidity (weight loss) of mice infected intraperitoneally (i.p.) with GAS 3, 5, and 10 d after the initial viral infection. Mortality was also significantly increased when mice were infected with GAS 3 and 5 d after pulmonary IAV infection. Increased mortality among mice infected with virus 5 d prior to bacterial infection correlated with increased dissemination of GAS from the peritoneum to the blood, spleen, and lungs. The interval was also associated with a significant increase in the pro-inflammatory cytokines IFN-γ, IL-12, TNF-α, MCP-1 and IL-27 in sera. We conclude, using a murine model, that respiratory influenza virus infection increases the likelihood and severity of systemic iGAS disease, even when GAS infection does not originate in the respiratory tract.

Epidemiological and Molecular Study on Tick-Borne Pathogens in Argun Port Area Near the Chinese-Russian Border.

Objective: We aim to investigate the species composition of ticks and the pathogen characteristics they carry in the Argun port area of the China-Russia border. Materials and Methods: Ticks were collected in surrounding grassland, mixed forest land, and other different habitats around the Argun port area at the Sino-Russian Border of Inner Mongolia in China in April 2019. The presence of 16 potential pathogens, including Yersinia Pestis, Francisella tularensis, Coxiella burnetii (Cb), Anaplasma sp. (Ap), spotted fever group rickettsiae (SFG Rk), Borrelia sp. (Bl), Leptospira, Bartonella spp., Babesia, Crimean-Congo hemorrhagic fever virus, tick-borne encephalitis virus, Bhanja virus, West Nile Virus, severe fever with thrombocytopenia syndrome bunyavirus, Hantaan virus, and bocavirus (boca) was analyzed by polymerase chain reaction. The DNA and amino acid sequences of tick-borne pathogens were compared for homology, and the phylogenetic trees were constructed by using Mega and Lasergene software. Results: A total of 210 ticks were collected and they belonged to three species: Dermacentor nuttalli, Ixodes persulcatus, and Haemaphysalis verticalis. Among them, 165 (78.57%) ticks tested positive for 5 pathogens, namely Ap, SFG Rk, Cb, Bl, and boca. Fifteen (7.14%) ticks were detected coinfection with two pathogens, and none were coinfected with three or more pathogens. Conclusion: This study shows the prevalence of at least five tick-borne pathogens in Argun, and there is a risk of coinfection by two pathogens in one tick. This study reveals the great importance of controlling tick-borne diseases in this region.

A meta-meta-analysis of co-infection, secondary infections, and antimicrobial resistance in COVID-19 patients.

The newly discovered coronavirus SARS-CoV-2 has sparked a worldwide pandemic of COVID-19, which has caused havoc on medical infrastructures, economies, and cultures around the world. Determining the whole scenario is essential since SARS-CoV-2 variants and sub-variants keep appearing after vaccinations and booster doses. The objective of this secondary meta-analysis is to analysis co-infection, secondary infections, and antimicrobial resistance (AMR) in COVID-19 patients. This study used five significant databases to conduct a systematic review and an overlap meta-analysis to evaluate the pooled estimates of co-infections and secondary infections. The summary of the meta-analysis showed an overall co-infection effect of 26.19% (95% confidence intervals CI: 21.39-31.01, I2 =98.78, n = 14 meta-analysis) among patients with COVID-19. A coinfection effect of 11.13% (95% CI: 9.7-12.56, I2 =99.14, n = 11 meta-analysis) for bacteria; 9.69% (95% CI: 1.21-7.90, I2 =98.33) for fungal and 3.48% (95% CI: 2.15-4.81, I2 =95.84) for viruses. A secondary infection effect of 19.03% (95% CI: 9.53-28.54, I2 =85.65) was pooled from 2 meta-analyses (Ave: 82 primary studies). This is the first study that compiles the results of all the previous three years meta-analyses into a single source and offers strong proof of co-infections and secondary infections in COVID-19 patients. Early detection of co-infection and AMR is crucial for COVID-19 patients in order to effective treatment.

Within- and between-host dynamics of producer and non-producer pathogens.

For infections to be maintained in a population, pathogens must compete to colonize hosts and transmit between them. We use an experimental approach to investigate within-and-between host dynamics using the pathogen Pseudomonas aeruginosa and the animal host Caenorhabditis elegans. Within-host interactions can involve the production of goods that are beneficial to all pathogens in the local environment but susceptible to exploitation by non-producers. We exposed the nematode host to 'producer' and two 'non-producer' bacterial strains (specifically for siderophore production and quorum sensing), in single infections and coinfections, to investigate within-host colonization. Subsequently, we introduced infected nematodes to pathogen-naive populations to allow natural transmission between hosts. We find that producer pathogens are consistently better at colonizing hosts and transmitting between them than non-producers during coinfection and single infection. Non-producers were poor at colonizing hosts and between-host transmission, even when coinfecting with producers. Understanding pathogen dynamics across these multiple levels will ultimately help us predict and control the spread of infections, as well as contribute to explanations for the persistence of cooperative genotypes in natural populations.

Analysis of coexisting pathogens in nasopharyngeal swabs from COVID-19.

Background: The impact of COVID-19 on the world is still ongoing, and it is currently under regular management. Although most infected people have flu-like symptoms and can cure themselves, coexisting pathogens in COVID-19 patients should not be taken lightly. The present study sought to investigate the coexisting pathogens in SARS-CoV-2 infected patients and identify the variety and abundance of dangerous microbes to guide treatment strategies with a better understanding of the untested factors. Methods: We extracted total DNA and RNA in COVID-19 patient specimens from nasopharyngeal swabs to construct a metagenomic library and utilize Next Generation Sequencing (NGS) to discover chief bacteria, fungi, and viruses in the body of patients. High-throughput sequencing data from Illumina Hiseq 4000 were analyzed using Krona taxonomic methodology for species diversity. Results: We studied 56 samples to detect SARS-CoV-2 and other pathogens and analyzed the species diversity and community composition of these samples after sequencing. Our results showed some threatening pathogens such as Mycoplasma pneumoniae, Klebsiella pneumoniae, Streptococcus pneumoniae, and some previously reported pathogens. SARS-CoV-2 combined with bacterial infection is more common. The results of heat map analysis showed that the abundance of bacteria was mostly more than 1000 and that of viruses was generally less than 500. The pathogens most likely to cause SARS-CoV-2 coinfection or superinfection include Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus aureus, Klebsiella pneumoniae, and Human gammaherpesvirus 4. Conclusions: The current coinfection and superinfection status is not optimistic. Bacteria are the major threat group that increases the risk of complications and death in COVID-19 patients and attention should be paid to the use and control of antibiotics. Our study investigated the main types of respiratory pathogens prone to coexisting or superinfection in COVID-19 patients, which is valuable for identifying and treating SARS-CoV-2.

Epidemiology of gastrointestinal infections: lessons learned from syndromic testing, Region Zealand, Denmark.

The aim of this study was to investigate the value of syndromic diagnostic testing for a better understanding of the epidemiology of gastrointestinal infections in Denmark. Here we evaluated the QIAstat-Dx® Gastrointestinal (GI) Panel 1 assay on 18,610 fecal samples requested for analysis for enteric pathogens in Region Zealand, Denmark, in 1 year (October 1, 2021, to September 30, 2022). In total, 6905 (37%) samples were detected positive for one or more diarrhoeal bacteria, viruses, and protozoa. The most common bacterial, viral, and parasitic pathogens detected with the QIAstat-Dx® Gastrointestinal Panel 1 were EPEC (in patients ≥ 2 years of age) (n = 1420 (20.6%)), rotavirus (n = 948 (13.7%)), and Cryptosporidium spp. (n = 196 (2.84%)). We identified a large diversity in infections likely reflecting substantial differences in the epidemiology including origin of infections, mode of transmission, seasonality, age-dependent susceptibility to disease, severity, and travel history. All pathogens were detected as both single and coinfections. Viral infections peaked in March with a positive rate of 31.6%, and bacterial infections peaked in August with a positive rate of 35.3%. ETEC, Shigella/EIEC, EAEC, and P. shigelloides were most related to travel activity, and coinfections were frequent. The distribution of Ct values varied significantly between the pathogens, with the lowest Ct values (median 17-18) observed in astrovirus, adenovirus, and rotavirus. Our results highlight the value of providing extensive diagnostic testing on fecal samples for sufficient detection of relevant diarrhoeal pathogens for optimal clinical care.

Multispecies coinfections and presence of antibiotics shape resistance and fitness costs in a pathogenic bacterium.

Increasing antimicrobial resistance (AMR) poses a challenge for treatment of bacterial diseases. In real life, bacterial infections are typically embedded within complex multispecies communities and influenced by the environment, which can shape costs and benefits of AMR. However, knowledge of such interactions and their implications for AMR in vivo is limited. To address this knowledge gap, we investigated fitness-related traits of a pathogenic bacterium (Flavobacterium columnare) in its fish host, capturing the effects of bacterial antibiotic resistance, coinfections between bacterial strains and metazoan parasites (fluke Diplostomum pseudospathaceum) and antibiotic exposure. We quantified real-time replication and virulence of sensitive and resistant bacteria and demonstrate that both bacteria can benefit from coinfection in terms of persistence and replication, depending on the coinfecting partner and antibiotic presence. We also show that antibiotics can benefit resistant bacteria by increasing bacterial replication under coinfection with flukes. These results emphasize the importance of diverse, inter-kingdom coinfection interactions and antibiotic exposure in shaping costs and benefits of AMR, supporting their role as significant contributors to spread and long-term persistence of resistance.

Altered Respiratory Microbiomes, Plasma Metabolites, and Immune Responses in Influenza A Virus and Methicillin-Resistant Staphylococcus aureus Coinfection.

Influenza A virus (IAV)-methicillin-resistant Staphylococcus aureus (MRSA) coinfection causes severe respiratory infections. The host microbiome plays an important role in respiratory tract infections. However, the relationships among the immune responses, metabolic characteristics, and respiratory microbial characteristics of IAV-MRSA coinfection have not been fully studied. We used specific-pathogen-free (SPF) C57BL/6N mice infected with IAV and MRSA to build a nonlethal model of IAV-MRSA coinfection and characterized the upper respiratory tract (URT) and lower respiratory tract (LRT) microbiomes at 4 and 13 days postinfection by full-length 16S rRNA gene sequencing. Immune response and plasma metabolism profile analyses were performed at 4 days postinfection by flow cytometry and liquid chromatography-tandem mass spectrometry (LC-MS/MS). The relationships among the LRT microbiota, the immune response, and the plasma metabolism profile were analyzed by Spearman's correlation analysis. IAV-MRSA coinfection showed significant weight loss and lung injury and significantly increased loads of IAV and MRSA in bronchoalveolar lavage fluid (BALF). Microbiome data showed that coinfection significantly increased the relative abundances of Enterococcus faecalis, Enterobacter hormaechei, Citrobacter freundii, and Klebsiella pneumoniae and decreased the relative abundances of Lactobacillus reuteri and Lactobacillus murinus. The percentages of CD4+/CD8+ T cells and B cells in the spleen; the levels of interleukin-9 (IL-9), interferon gamma (IFN-γ), tumor necrosis factor alpha (TNF-α), IL-6, and IL-8 in the lung; and the level of mevalonolactone in plasma were increased in IAV-MRSA-coinfected mice. L. murinus was positively correlated with lung macrophages and natural killer (NK) cells, negatively correlated with spleen B cells and CD4+/CD8+ T cells, and correlated with multiple plasma metabolites. Future research is needed to clarify whether L. murinus mediates or alters the severity of IAV-MRSA coinfection. IMPORTANCE The respiratory microbiome plays an important role in respiratory tract infections. In this study, we characterized the URT and LRT microbiota, the host immune response, and plasma metabolic profiles during IAV-MRSA coinfection and evaluated their correlations. We observed that IAV-MRSA coinfection induced severe lung injury and dysregulated host immunity and plasma metabolic profiles, as evidenced by the aggravation of lung pathological damage, the reduction of innate immune cells, the strong adaptation of the immune response, and the upregulation of mevalonolactone in plasma. L. murinus was strongly correlated with immune cells and plasma metabolites. Our findings contribute to a better understanding of the role of the host microbiome in respiratory tract infections and identified a key bacterial species, L. murinus, that may provide important references for the development of probiotic therapies.

Pleural effusion-related Nocardia otitidiscaviarum, Anaplasma platys and Ehrlichia canis coinfection in a dog.

The coinfections by some microorganisms have been related to severe diseases in humans and animals, where immunosuppressive agents favor opportunistic behavior of other pathogens. A 4-month-old, female mixed-breed dog with a two-week history of inappetence, prostration, emaciation, and respiratory distress was admitted at a veterinary hospital in Brazil. Tachycardia, pale mucous membranes, severe respiratory distress, and a large number of ticks (Rhipicephalus sanguineus s.l.) in different body regions were observed at clinical examination. Hematological examination of dog showed leukocytosis, neutrophilia, mild anemia, and thrombocytopenia, whereas unremarkable values in biochemical tests. Thoracic radiography revealed a pleural effusion image. Blood and the pleural fluid (purulent aspect) samples were subjected to qPCR (16S rRNA and dsb genes) and sequencing, which identified Ehrlichia canis and Anaplasma platys coinfection. An aggregate of coccoid-to-branching or long filamentous microorganisms, surrounded by pyogranulomatous inflammatory reaction was seen at the cytology of the pleural fluid. Bacteriological culture of pleural effusion showed colonies compatible with the genus Nocardia, which revealed gram-positive filamentous organisms with a tendency of fragmentation and were identified as Nocardia otitidiscaviarum in mass spectrometry (MALDI-TOF MS). Therapy of N. otitidiscaviarum isolate using levofloxacin (supported by a previous in vitro susceptibility testing) and doxycycline for E. canis and A. platys resulted in complete resolution of the clinical picture. Here, we report for the first time a triple coinfection by Nocardia otitidiscaviarum, A. platys, and E. canis in a dog with pleural effusion, where debilitating or immunosuppressive conditions induced by A. platys and E. canis coinfection probably contributed to the opportunistic behavior of N. otitidiscaviarum.

Characterisation of key genotypic and phenotypic traits of clinical cystic fibrosis Staphylococcus aureus isolates.

Introduction. One third of people with CF in the UK are co-infected by both Staphylococcus aureus and Pseudomonas aeruginosa. Chronic bacterial infection in CF contributes to the gradual destruction of lung tissue, and eventually respiratory failure in this group.Gap Statement. The contribution of S. aureus to cystic fibrosis (CF) lung decline in the presence or absence of P. aeruginosa is unclear. Defining the molecular and phenotypic characteristics of a range of S. aureus clinical isolates will help further understand its pathogenic capabilities.Aim. Our objective was to use molecular and phenotypic tools to characterise twenty-five clinical S. aureus isolates collected from mono- and coinfection with P. aeruginosa from people with CF at the Royal Victoria Infirmary, Newcastle upon Tyne.Methodology. Genomic DNA was extracted and sequenced. Multilocus sequence typing was used to construct phylogeny from the seven housekeeping genes. A pangenome was calculated using Roary, and cluster of Orthologous groups were assigned using eggNOG-mapper which were used to determine differences within core, accessory, and unique genomes. Characterisation of sequence type, clonal complex, agr and spa types was carried out using PubMLST, eBURST, AgrVATE and spaTyper, respectively. Antibiotic resistance was determined using Kirby-Bauer disc diffusion tests. Phenotypic testing of haemolysis was carried out using ovine red blood cell agar plates and mucoid phenotypes visualised using Congo red agar.Results. Clinical strains clustered closely based on agr type, sequence type and clonal complex. COG analysis revealed statistically significant enrichment of COG families between core, accessory and unique pangenome groups. The unique genome was significantly enriched for replication, recombination and repair, and defence mechanisms. The presence of known virulence genes and toxins were high within this group, and unique genes were identified in 11 strains. Strains which were isolated from the same patient all surpassed average nucleotide identity thresholds, however, differed in phenotypic traits. Antimicrobial resistance to macrolides was significantly higher in the coinfection group.Conclusion. There is huge variation in genetic and phenotypic capabilities of S. aureus strains. Further studies on how these may differ in relation to other species in the CF lung may give insight into inter-species interactions.