Influenza poses a serious threat to both individual and public health. This study aimed to investigate the virological and epidemiological characteristics of influenza infections and to explore the genetic diversity of the circulating influenza viruses. In total, 1886 nasopharyngeal specimens from patients with acute respiratory illnesses were tested against 13 respiratory viruses using a multiplex real-time PCR. Whole-genome sequencing, phylogenetic, and amino acid analyses of representative influenza strains were performed. At least one respiratory virus was detected in 869 (46.1%) patients; 87 (4.6%) were co-infected with two or three viruses. Influenza A(H1N1)pdm09 was the most prevalent virus (16.1%), followed by rhinoviruses (8.1%) and RSV (6.7%). Hemagglutinin (HA) genes of the 74 influenza A(H1N1)pdm09 viruses were categorized in subclades C.1.8, C.1.9, and C.1 within clade 5a.2a and D1, D.2, and D.3 within clade 5a.2a.1. The A(H3N2) viruses analyzed belonged to clade 2a.3a.1, subclades J.2 and J.1. The sequenced B/Victoria lineage viruses fell into clade V1A.3a.2, subclades C.5.6 and C.5.7. Amino acid substitutions in most viral proteins were identified compared with the vaccine strains, including in the HA antigenic sites. This study demonstrated the dominant distribution of the influenza A(H1N1)pdm09 virus among the respiratory viruses studied and the genetic diversity of the circulating influenza viruses.

The increased life expectancy of PLHIV (People Living with HIV) and the successful highly combined antiretroviral therapy (cART) poses new clinical challenges regarding aging and its co-morbid condition. It is commonly believed that HIV infection “accelerates” aging. Human immunodeficiency virus type 1 (HIV-1) infection is characterized by inflammation and immune activation that persists despite cART, and that may contribute to the development of co-morbid conditions. In this regard, we aimed to compare current cART regimens in light of premature aging to evaluate differences in their ability to reduce immune activation and inflammation in virologically suppressed patients. We studied a panel of biomarkers (IFN-γ, IL-1β, IL-12p70, IL-2, IL-4, IL-5, IL-6, IL-13, IL-18, GM-CSF, TNF-α, C-reactive protein, D-dimer, soluble CD14), which could provide a non-invasive and affordable approach to monitor HIV-related chronic inflammation. The results of the current study do not provide hard evidence favoring a particular cART regimen, although they show a less favorable regimen profile containing a protease inhibitor. Our data suggest an incomplete reduction of inflammation and immune activation in terms of the effective cART. It is likely that the interest in various biomarkers related to immune activation and inflammation as predictors of clinical outcomes among PLHIV will increase in the future.

Background: Mixed infections can worsen disease symptoms. This study investigated the impact of mixed infections with viral and bacterial pathogens in patients positive for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Methods: Using the in-house multiplex PCR method, we tested 337 SARS-CoV-2 positive samples for co-infections with three bacterial and 14 other viral pathogens. Results: Between August 2021 and May 2022, 8% of 337 SARS-CoV-2-positive patients had bacterial co-infections, 5.6% had viral co-infections, and 1.4% had triple mixed infections. The most common causes of mixed infections were Haemophilus influenzae (5.93%) and respiratory syncytial virus (RSV) (1.18%). Children < 5 years old had more frequent co-infections than adults < 65 years old (20.8% vs. 16.4%), while adults showed a more severe clinical picture with a higher C-reactive protein (CRP) level (78.1 vs.16.2 mg/L; p = 0.033), a lower oxygen saturation (SpO2) (89.5 vs. 93.2%), and a longer hospital stay (8.1 vs. 3.1 days; p = 0.025) (mean levels). The risk of a fatal outcome was 41% in unvaccinated patients (p = 0.713), which increased by 2.66% with co-infection with two pathogens (p = 0.342) and by 26% with three pathogens (p = 0.005). Additionally, 50% of intensive care unit (ICU) patients had a triple infection, compared with only 1.3% in the inpatient unit (p = 0.0029). The risk of death and/or ICU admission was 12 times higher (p = 0.042) with an additional pathogen and increased by 95% (p = 0.003) with a third concomitant pathogen. Conclusions: Regular multiplex testing is important for prompt treatment and targeted antibiotic use.

Background/Objectives: Monkeypox (mpox) is currently the most common orthopoxvirus (OPXV) zoonotic disease, and, since 2022, there has been atypical person-to-person transmission observed in non-endemic countries. The present study aimed to investigate the frequency of monkeypox virus (MPXV) and OPXV DNA detection in recommended and alternative clinical materials taken during the acute and convalescent phases of infection in Bulgarian patients. Methods: The study included laboratory investigation by real time PCR of 181 clinical samples from 42 Bulgarian patients with possible mpox infections. Results: MPXV DNA was detected in 23/181 (12.71%), and OPXV DNA in 20/181 (11.05%) clinical samples. There were six mpox-confirmed patients aged 23 to 44. At the highest frequency, MPXV and OPXV DNA were detected in samples of vesicular contents (6/6) and nasal/oropharyngeal secretions (5/6 and 4/6) during the first three days from the appearance of clinical symptoms. We demonstrated MPXV and OPXV DNA in alternative samples (urine, feces, ejaculate, and saliva), and in follow-up patient samples, taken two weeks after mpox confirmation in the convalescent phase (vesicular contentsand urine). Conclusions: Our findings suggested that MPXV may be detected in a larger set of clinical materials, including alternatives, where the virus can persist for more than two weeks.

Monkeypox is an emerging zoonosis that has been declared a public health threat due to its spread outside of endemic areas. We present two cases of monkeypox: one involving a 39-year-old male patient living with HIV on antiretroviral therapy with detectable viraemia and a low CD4 count, and another involving a 44-year-old male, negative for HIV, who visited our Diagnostic and Consultative Unit with a new-onset skin rash. Both individuals identify as men who have sex with men and had travelled abroad before the disease onset. One patient was hospitalised, while the other was treated as an outpatient with follow-up care. Both cases were managed conservatively due to the unavailability of tecovirimat in Bulgaria. We emphasise the importance of recognising and diagnosing monkeypox and the significance of HIV screening, as monkeypox could be considered an AIDS-defining condition in people living with HIV. Maintaining a high level of suspicion for new cases is essential, as instances of monkeypox continue to be reported in non-endemic countries.

Background: SARS-CoV-2 can trigger hyperinflammation, leading to severe COVID-19, presenting with pneumonia, acute respiratory distress syndrome (ARDS), and multiple organ failure. Specific biomarkers like leukocytes, CRP, NLR, AST, LDH, D-dimer, ferritin, and IL-6 are associated with disease severity. Anakinra, an IL-1 receptor antagonist, has been proposed to mitigate hyperinflammation, but its clinical efficacy remains uncertain. This study aimed to evaluate the effect of Anakinra on inflammatory biomarkers, oxygenation status, and survival outcomes in hospitalized patients with moderate to severe COVID-19 (according to the National Institute of Health severity scale), compared to standard treatment. Methods: A retrospective analysis included 65 patients (mean age 75.51 ± 9.54 years; 58.5% male, 41.5% female) hospitalized with moderate to severe COVID-19. Patients were divided into two groups: a control group receiving standard treatment (n = 24) and a target group treated with Anakinra (n = 41). Biomarkers and oxygenation status were assessed on Days 0, 3, and 7. Statistical analyses compared the groups for changes in leukocytes, NLR, CRP, AST, LDH, D-dimer, ferritin, and IL-6. Results: Anakinra treatment was associated with significant reductions in leukocytes, NLR, D-dimer, ferritin, IL-6, and CRP by Days 3 and 7. Improvements in oxygenation status were observed, although no survival benefits were noted. The control group showed no significant biomarker changes except for AST and LDH on Day 7. Conclusions: Anakinra demonstrated favorable effects on biomarkers and oxygenation in moderate to severe COVID-19 but did not improve survival. Further studies are needed to validate these findings.