Numerous protocols for viral enrichment and genome amplification have been created. However, the direct identification of viral genomes from clinical specimens using next-generation sequencing (NGS) still has its challenges. As a selected viral nucleic acid extraction method may determine the sensitivity and reliability of NGS, it is still valuable to evaluate the extraction efficiency of different extraction kits using clinical specimens directly. In this study, we performed qRT-PCR and viral metagenomic analysis of the extraction efficiency of four commonly used Qiagen extraction kits: QIAamp Viral RNA Mini Kit (VRMK), QIAamp MinElute Virus Spin Kit (MVSK), RNeasy Mini Kit (RMK), and RNeasy Plus Micro Kit (RPMK), using a mixed respiratory clinical sample without any pre-treatment. This sample contained an adenovirus (ADV), influenza virus A (Flu A), human parainfluenza virus 3 (PIV3), human coronavirus OC43 (OC43), and human metapneumovirus (HMPV). The quantity and quality of the viral extracts were significantly different among these kits. The highest threshold cycle(Ct)values for ADV and OC43 were obtained by using the RPMK. The MVSK had the lowest Ct values for ADV and PIV3. The RMK revealed the lowest detectability for HMPV and PIV3. The most effective rate of NGS data at 67.47% was observed with the RPMK. The other three kits ranged between 12.1–26.79% effectiveness rates for the NGS data. Most importantly, compared to the other three kits the highest proportion of non-host reads was obtained by the RPMK. The MVSK performed best with the lowest Ct value of 20.5 in the extraction of ADV, while the RMK revealed the best extraction efficiency by NGS analysis. The evaluation of viral nucleic acid extraction efficiency is different between NGS and qRT-PCR analysis. The RPMK was most applicable for the metagenomic analysis of viral RNA and enabled more sensitive identification of the RNA virus genome in respiratory clinical samples. In addition, viral RNA extraction kits were also applicable for metagenomic analysis of the DNA virus. Our results highlighted the importance of nucleic acid extraction kit selection, which has a major impact on the yield and number of viral reads by NGS analysis. Therefore, the choice of extraction method for a given viral pathogen needs to be carefully considered.

Wohlfahrtiimonas chitiniclastica is a rare Gram-negative bacterium typically associated with wound infections, particularly in immunocompromised patients or individuals exposed to unsanitary conditions. Although clinical cases are infrequent, the infection can lead to severe complications such as bacteremia, septic shock, and even death if unrecognized or inadequately treated. We present the case of a 76-year-old diabetic male who developed chronic, non-healing foot ulcers complicated by gangrene and maggot infestation. Diagnostic challenges were addressed using metagenomic next-generation sequencing, which identified W. chitiniclastica alongside Proteus mirabilis and Corynebacterium striatum. Management included aggressive surgical debridement to remove necrotic tissue, local application of gentamicin-impregnated bone cement, systemic antibiotic therapy with ertapenem, and wound reconstruction using a dorsally based fascial flap. This combined approach resulted in significant clinical improvement, progressive wound healing, and marked reductions in infection markers. The case highlights the decisive role of advanced sequencing technologies in identifying rare pathogens within polymicrobial infections, where conventional methods such as MALDI-TOF mass spectrometry may fail. It also emphasizes the importance of integrating precision diagnostics with surgical intervention, targeted antimicrobial therapy, and rigorous postoperative care to achieve successful outcomes. By documenting this unusual presentation, we aim to expand clinical awareness of W. chitiniclastica infections and provide a practical framework for managing similarly complex diabetic foot infections.

Wohlfahrtiimonas chitiniclastica is a rare Gram-negative bacterium typically associated with wound infections, particularly in immunocompromised patients or individuals exposed to unsanitary conditions. Although clinical cases are infrequent, the infection can lead to severe complications such as bacteremia, septic shock, and even death if unrecognized or inadequately treated. We present the case of a 76-year-old diabetic male who developed chronic, non-healing foot ulcers complicated by gangrene and maggot infestation. Diagnostic challenges were addressed using metagenomic next-generation sequencing, which identified W. chitiniclastica alongside Proteus mirabilis and Corynebacterium striatum. Management included aggressive surgical debridement to remove necrotic tissue, local application of gentamicin-impregnated bone cement, systemic antibiotic therapy with ertapenem, and wound reconstruction using a dorsally based fascial flap. This combined approach resulted in significant clinical improvement, progressive wound healing, and marked reductions in infection markers. The case highlights the decisive role of advanced sequencing technologies in identifying rare pathogens within polymicrobial infections, where conventional methods such as MALDI-TOF mass spectrometry may fail. It also emphasizes the importance of integrating precision diagnostics with surgical intervention, targeted antimicrobial therapy, and rigorous postoperative care to achieve successful outcomes. By documenting this unusual presentation, we aim to expand clinical awareness of W. chitiniclastica infections and provide a practical framework for managing similarly complex diabetic foot infections.