Discover the three studies we selected in April’s Microbiology Time.

• The first study we selected explored the potential of using tongue swabbing as a substitute for sputum in tuberculosis (TB) testing. Previous research indicated that the Cepheid Xpert Ultra platform performs well compared to reference standards on buffer-preserved sputum samples. Here, the researchers assessed the same platform on 245 self-collected and dry-stored tongue swabs and compared the results with the Molbio Truenat MTB Ultima (MTB Ultima) assay. Xpert Ultra showed 75.5% sensitivity and 100% specificity compared to sputum testing, while MTB Ultima demonstrated 71.6% sensitivity and 96.9% specificity, improving to 79.1% sensitivity after re-testing frozen samples initially yielding false-negative or invalid results. These findings highlight the feasibility of using tongue swabs as easy-to-collect samples for TB testing.
• For the second study, we move to Indonesia, where Dr. Wulandri and colleagues investigated the pathogens causing acute diarrhea in children, comparing HIV-infected and non-HIV-infected cases at an Indonesian tertiary hospital. Conducted from March 2019 to April 2020, the cross-sectional study utilized multiplex RT-PCR, showing that almost 60% of the samples were positive, with more positive results in HIV-infected children than in non-HIV-infected children (70% vs. 54.7%). Overall, 72 enteropathogens were detected, with non-HIV cases primarily showing bacterial infections with enteroaggregative Escherichia coli as predominant, followed by viral infections. In contrast, HIV-infected children exhibited a mix of viruses, bacteria, and parasites with Cryptosporidium spp. solely detected in this group. Notably, non-HIV children predominantly presented with invasive bacterial pathogens, while HIV-infected children showed more viral and parasitic infections, often attributed to opportunistic pathogens.
• The last study investigated the relationship between gut microbiome composition and clinical outcomes in COVID-19 patients. Stool samples and patient records were collected from 24 hospitalized individuals with confirmed SARS-CoV-2 infection. Fungal and bacterial gut microbiome was characterized by amplicon sequencing on the Illumina MiSeq. The results revealed that female COVID-19 patients had lower gut bacterial microbiota richness compared to males, possibly influenced by differences in hospital admission timing. Both sexes exhibited associations with opportunistic bacterial pathogens like Enterococcus and Streptococcus. The Candida genus dominated the gut mycobiota of COVID-19 patients, with adults showing higher fungal diversity than the elderly. Notably, Saccharomycetales unassigned fungal genera correlated with bacterial short-chain fatty acid producers and negatively with proinflammatory bacteria like Bilophila. Patients harboring this fungal taxon were not admitted to high-intensity units. The study underscores the association between COVID-19 and opportunistic bacterial pathogens, highlights Candida dominance in the gut, and suggests a potential protective role for specific fungal taxa, warranting further investigation into their role in gastrointestinal health and disease outcomes.

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This month, we bring you a new oral microbiota sampling method, a study on saliva and Long COVID, and yet another paper highlighting the importance of self-collection in the fight against cervical cancer.

• This first study investigated the association between COVID-19 and Long COVID in children. The Italian researchers conducted a prospective study involving 167 pediatric patients hospitalized with confirmed acute SARS-CoV-2 infection, and collecting saliva samples with LolliSponge. A telephone survey was conducted at 3-6-12 months after discharge. Of the patients, 95.5% presented with symptoms upon admission. Regarding patients for whom it was feasible to determine the SARS-CoV-2 variant, the study identified the Delta variant in 21.6% and the Omicron variant in 78.4%. About 16.5% of patients reported symptoms indicative of Long COVID, such as weight loss, inappetence, chronic cough, fatigue, and others. Only 4 patients with Long COVID had a specific SARS-CoV-2 variant identified. The study emphasizes the significant concern of Long COVID in the pediatric population, highlighting the need for continuous monitoring and tailored symptom investigation based on age.
• For the second study, we remain in Italy. Marianna Martinelli and colleagues assessed the accuracy of the BD Onclarity HPV assay on self-collected vaginal FLOQSwabs compared to cervical samples for detecting cervical intraepithelial neoplasia (CIN). Testing was conducted on two automated platforms, BD Viper LT and BD COR, and included 300 women. Self-sampling was performed with two vaginal swabs to assess, with the second one, the influence of freezing on the results. The results showed similar sensitivity for ≥CIN2 between vaginal and cervical samples, regardless of the platform used. Specificity was initially slightly lower for self-collected samples compared to cervical samples, but cut-off optimization for HPV positivity improved relative specificity without compromising sensitivity. No significant differences were observed between self-samples tested with the two platforms and between first- and second-collected (frozen) swabs. The study concludes that Onclarity assay on self-collected FLOQSwabs samples showed similar sensitivity and specificity (with cut-off optimization) to detect ≥CIN2 compared to cervical samples.
• The third study addressed the challenge of standardizing oral microbiome analysis due to the variability in microbiota compositions across different oral habitats and the lack of standardized collection methods. The study proposes a standardized protocol for dental examination and material collection for next-generation sequencing (NGS) from various oral sites. The researchers involved 60 patients grouped into COVID-19 convalescents with and without antibiotics and healthy individuals. The collected biological samples were then used to conduct NGS (16S rRNA). Alpha and beta diversity analyses reveal notable differences in microbial communities, particularly in the supragingival plaque. The study emphasizes the importance of precise procedures during dental exams and material collection to avoid technical errors and contamination in NGS results. Dental indices like DMFT, D number, PI, and BOP are highlighted as valuable prognostic parameters for assessing oral health. The developed procedures are suggested for use in future studies by future studies.

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