Mycoplasma genitalium Antimicrobial Resistance in Community and Sexual Health Clinic Patients, Auckland, New Zealand.

Our retrospective study compared genotypic antimicrobial resistance in Mycoplasma genitalium-positive specimens collected from 48 community and 33 sexual health clinic (SHC) patients. Macrolide resistance was similar in community (75%) and SHC (76%) patients. We observed no significant difference in fluoroquinolone resistance between community (19%) and SHC (27%) patients (p = 0.66).

Oral microbial influences on oral mucositis during radiotherapy treatment of head and neck cancer.

PURPOSE: Oral mucositis (OM) remains a significant complication developed by many patients undergoing radiotherapy (RT) to the head and neck region. Emerging data suggest oral microbes may contribute to the onset and severity of this acute side effect. METHODS: In this study, saliva and oral swabs from head and neck cancer patients undergoing RT were collected. We employed molecular microbiological techniques to study the bacterial communities present in saliva, and both the bacterial and fungal communities present on the buccal mucosa and lateral tongue. Changes in microbiota composition with increasing radiation dose and the presence of mucositis were examined. RESULTS: The data suggest that the salivary microbiota remain stable during RT and are consistently dominated by Streptococcus, Prevotella, Fusobacterium and Granulicatella. Obligate and facultative anaerobic Gram-negative bacilli (GNB) Bacteroidales G2, Capnocytophaga, Eikenella, Mycoplasma and Sneathia, as well as anaerobic GNB in the periopathogenic genera Porphyromonas and Tannerella, were all positively correlated with ≥ grade 2 OM. Significant increases in the relative abundances of Bacteroidales G2, Fusobacterium and Sneathia were identified in buccal mucosa swabs at sites of ≥ grade 2 OM (p < 0.05). Furthermore, the abundance of several GNB (Fusobacterium, Haemophilus, Tannerella, Porphyromonas and Eikenella) on the buccal mucosa may influence patient susceptibility to developing OM. Candida was widely detected in buccal mucosa swabs, regardless of mucositis status. CONCLUSIONS: Our findings support previously hypothesized associations between oral health and the pathogenesis of OM, highlighting the importance of oral health interventions for head and neck cancer patients.

Evaluation of ssrA-targeted real time PCR for the detection of Bartonella species in human clinical samples and reflex sequencing for species-level identification.

The genus Bartonella includes species capable of causing disease in animals and humans. Due to its fastidious nature, direct detection of Bartonella causing human infection relies largely on molecular microbiological methods. Thus, it is imperative that diagnostic assays in use have the ability to detect a range of Bartonella species associated with human disease. In this study, we compared the performance of a real time polymerase chain reaction (PCR) assay targeting the ssrA gene to conventional rpoB-targeted PCR and sequencing for detection and differentiation of Bartonella species in human clinical samples. The real time ssrA PCR performed better for non-Bartonella henselae species, detecting B. clarridgeiae and B. quintana DNA in heart valve specimens that were not detected by rpoB PCR, and improved the sensitivity of B. henselae detection in blood specimens. Our findings suggest the real time ssrA PCR assay is suitable for detection and identification of Bartonella species in human clinical specimens.

The emergence of azole resistance in Aspergillus fumigatus complex in New Zealand.

BACKGROUND: Azole resistance in Aspergillus fumigatus (A. fumigatus) is increasing globally. A pan-azole-resistant isolate prompted genetic analysis of local azole-resistant isolates to determine resistance genotypes. METHODS: All A. fumigatus complex isolates were tested by the broth colorimetric micro-dilution method, Sensititre® YeastOne® (SYO) (TREK Diagnostic Systems, West Sussex, England). Epidemiological cutoff values derived from the Clinical & Laboratory Standards Institute method were used to determine the proportion of non-wild-type (non-WT) isolates (ie, those with an increased likelihood to harbour acquired mechanisms of resistance). Non-WT isolates were identified by ß-tubulin gene sequencing and the genotype for azole resistance was determined. The history of the patient with the first pan-resistant isolate was reviewed along with the treatment history of patients with azole-resistant strains. RESULTS: From January 2001 to August 2020, antifungal susceptibility testing was performed on 260 A. fumigatus complex isolates: six isolates were non-WT for one or more azole agent, two A. fumigatus sensu stricto and four other members within the species complex: two A. fischeri and two A. lentulus. There were three non-WT isolates for amphotericin B, three for itraconazole, five for posaconazole and five for voriconazole. All six non-WT strains were isolated in the past nine years (P<0.01), and four in the past three years. Azole-resistance genotyping for the A. fumigatus sensu stricto isolates detected amino acid changes at hot spots in the cyp51A gene: one at G54E and one at G138C. All six isolates were WT for caspofungin. Five of the six patients with azole-resistant strains had previous azole treatment, and the patient with the pan-azole-resistant strain had been on continuous azole treatment for 42 months preceding strain isolation. CONCLUSIONS: New Zealand can be added to the growing list of countries with azole-resistant A. fumigatus complex isolates, including pan-azole resistance in A. fumigatus sensu stricto. While uncommon and mostly found in cryptic species within the complex, azole resistance is increasing. The results provide a baseline for monitoring this emerging antifungal resistance trend in A. fumigatus in New Zealand.

Randomised, double-blind, placebo-controlled trial of oral probiotic Streptococcus salivarius M18 on head and neck cancer patients post-radiotherapy: a pilot study.

Xerostomia detrimentally affects the oral health of many head and neck cancer patients who undergo radiotherapy. Its sequelae become an ongoing burden for patients that often manifest as periodontal disease and dental decay. Bacteria play a major role in the pathogenesis of these conditions and here we explore the use of an oral probiotic to beneficially modulate the oral bacterial community post-radiotherapy. In this pilot study, a four-week intervention with oral probiotic lozenges containing Streptococcus salivarius M18 was trialled in seven patients. Post-intervention changes in oral health and in the composition of the plaque and saliva bacterial communities were compared with six patients in a placebo group. An improvement in periodontal screening and plaque index scores was observed in both groups after the intervention period. The oral probiotic lozenges did not significantly impact bacterial community composition or diversity, nor did the probiotic lozenges increase the relative sequence abundance of ZOTU_1 (the probiotic-associated sequence assigned to S. salivarius) detected in the samples. Network analyses suggest negative interactions occurred between ZOTU_1 and species from the periopathogenic genera Campylobacter, Fretibacterium, Selenomonas and Treponema but further investigation is required to more fully understand the beneficial properties of this oral probiotic.

Helicobacter cinaedi bacteremia in a returning traveler.

Of the non-Helicobacter pylori Helicobacter (NHPH) species, Helicobacter cinaedi is an emerging cause of infection in humans. Here we report a novel clinical presentation of H. cinaedi infection: a case of fever in a returning traveler. A 31 year old previously fit and well male presented with onset of fever 24 h after returning from travel in Singapore and Indonesia. Associated symptoms consisted of sore throat, mild shortness of breath, generalized myalgia and arthralgia, headache, and four episodes of loose stools. The patient recovered spontaneously without treatment and was discharged. After 4 days of incubation, blood cultures grew H. cinaedi. H. cinaedi is a slow-growing fastidious organism poorly detected by some commonly used automated blood culture systems, and difficult to identify using commercial or traditional biochemical identification systems. This case illustrates the importance of H. cinaedi as an emerging pathogen in immunocompetent patients, with a wide variety of possible clinical presentations. The challenges in the microbiological diagnosis of H. cinaedi infections lead us to speculate that H. cinaedi is an underdiagnosed cause of febrile illness, both in returning travelers and in other clinical settings.

Characterizing the Human Mycobiota: A Comparison of Small Subunit rRNA, ITS1, ITS2, and Large Subunit rRNA Genomic Targets.

Interest in the human microbiome has increased dramatically in the last decade. However, much of this research has focused on bacteria, while the composition and roles of their fungal counterparts remain less understood. Furthermore, a variety of methodological approaches have been applied, and the comparability between studies is unclear. This study compared four primer pairs targeting the small subunit (SSU) rRNA (18S), ITS1, ITS2, and large subunit (LSU) rRNA (26S) genomic regions for their ability to accurately characterize fungal communities typical of the human mycobiota. All four target regions of 21 individual fungal mock community taxa were capable of being amplified adequately and sequenced. Mixed mock community analyses revealed marked variability in the ability of each primer pair to accurately characterize a complex community. ITS target regions outperformed LSU and SSU. Of the ITS regions, ITS1 failed to generate sequences for Yarrowia lipolytica and all three Malassezia species when in a mixed community. These findings were further supported in studies of human sinonasal and mouse fecal samples. Based on these analyses, previous studies using ITS1, SSU, or LSU markers may omit key taxa that are identified by the ITS2 marker. Of methods commonly used in human mycobiota studies to date, we recommend selection of the ITS2 marker. Further investigation of more recently developed fungal primer options will be essential to ultimately determine the optimal methodological approach by which future human mycobiota studies ought to be standardized.

Microbial and inflammatory-based salivary biomarkers of head and neck squamous cell carcinoma.

Head and neck squamous cell carcinoma (HNSCC) patients often present with poor oral health, making it difficult to assess the relationship between oral microbes, inflammation, and carcinoma. This study investigates salivary microbes and inflammatory cytokines as biomarkers for HNSCC, with consideration of oral health. Saliva was collected from 30 participants, including 14 HNSCC patients and 16 participants representing both dentally compromised and healthy individuals. Bacterial and fungal communities were analyzed based on 16S rRNA gene and ITS1 amplicon sequencing, respectively, and concentrations of inflammatory cytokines were quantified using a cytometric bead array, with flow cytometry. Diversity-based analyses revealed that the bacterial communities of HNSCC patients were significantly different to those of the healthy control group but not the dentally compromised patients. Fungal communities were dominated by Candida, irrespective of cohort, with Candida albicans comprising ≥96% of fungal sequences in most HNSCC patients. Significantly higher concentrations of interleukin (IL)-1ß and IL-8 were detected in HNSCC and dentally compromised patients, when independently compared with healthy controls. IL-1ß and IL-8 concentrations were significantly positively correlated with the abundance of C. albicans. Our findings suggest that salivary microbial and inflammatory biomarkers of HNSCC are influenced by oral health.

Evaluating the Impact of DNA Extraction Method on the Representation of Human Oral Bacterial and Fungal Communities.

The application of high-throughput, next-generation sequencing technologies has greatly improved our understanding of the human oral microbiome. While deciphering this diverse microbial community using such approaches is more accurate than traditional culture-based methods, experimental bias introduced during critical steps such as DNA extraction may compromise the results obtained. Here, we systematically evaluate four commonly used microbial DNA extraction methods (MoBio PowerSoil® DNA Isolation Kit, QIAamp® DNA Mini Kit, Zymo Bacterial/Fungal DNA Mini PrepTM, phenol:chloroform-based DNA isolation) based on the following criteria: DNA quality and yield, and microbial community structure based on Illumina amplicon sequencing of the V3-V4 region of the 16S rRNA gene of bacteria and the internal transcribed spacer (ITS) 1 region of fungi. Our results indicate that DNA quality and yield varied significantly with DNA extraction method. Representation of bacterial genera in plaque and saliva samples did not significantly differ across DNA extraction methods and DNA extraction method showed no effect on the recovery of fungal genera from plaque. By contrast, fungal diversity from saliva was affected by DNA extraction method, suggesting that not all protocols are suitable to study the salivary mycobiome.