gyrA Mutations in Mycoplasma genitalium and Their Contribution to Moxifloxacin Failure: Time for the Next Generation of Resistance-Guided Therapy.

BACKGROUND: Although single nucleotide polymorphisms (SNPs) in Mycoplasma genitalium parC contribute to fluoroquinolone treatment failure, data are limited for the homologous gene, gyrA. This study investigated the prevalence of gyrA SNPs and their contribution to fluoroquinolone failure. METHODS: Samples from 411 patients (male and female) undergoing treatment for M. genitalium infection (Melbourne Sexual Health Centre, March 2019-February 2020) were analyzed by Sanger sequencing (gyrA and parC). For patients treated with moxifloxacin (n = 194), the association between SNPs and microbiologic treatment outcome was analyzed. RESULTS: The most common parC SNP was G248T/S83I (21.1% of samples), followed by D87N (2.3%). The most common gyrA SNP was G285A/M95I (7.1%). Dual parC/gyrA SNPs were found in 8.6% of cases. One third of infections harboring parC G248T/S83I SNP had a concurrent SNP in gyrA conferring M95I. SNPs in gyrA cooccurred with parC S83I variations. Treatment failure was higher in patients with parC S83I/gyrA dual SNPs when compared with infections with single S83I SNP alone from analysis of (1) 194 cases in this study (81.2% vs 45.8%, P = .047), and (2) pooled analysis of a larger population of 535 cases (80.6% vs 43.2%; P = .0027), indicating a strong additive effect. CONCLUSIONS: Compared with parC S83I SNP alone, M. genitalium infections with dual mutations affecting parC/gyrA had twice the likelihood of failing moxifloxacin. Although antimicrobial resistance varies by region globally, these data indicate that gyrA should be considered as a target for future resistance assays in Australasia. We propose a strategy for the next generation of resistance-guided therapy incorporating parC and gyrA testing.

Combination Therapy for Mycoplasma genitalium, and New Insights Into the Utility of parC Mutant Detection to Improve Cure.

BACKGROUND: Mycoplasma genitalium (MG) infection is challenging to cure because of rising antimicrobial resistance and limited treatment options. METHODS: This was a prospective evaluation of the efficacy and tolerability of resistance-guided combination antimicrobial therapy for MG treatment at Melbourne Sexual Health Centre (August 2019-December 2020). All patients received 7 days of doxycycline before combination therapy based on the macrolide-resistant profile. Macrolide-susceptible infections received combination doxycycline + azithromycin (1 g, day 1; 500 mg, days 2-4) and macrolide-resistant infections combination doxycycline + moxifloxacin (400 mg daily for 7 days). Adherence and adverse effects were recorded at test-of-cure, recommended 14-28 days after antimicrobial completion. Sequencing was performed to determine the prevalence of single nucleotide polymorphisms (SNPs) in the parC gene and their association with moxifloxacin treatment outcomes in macrolide-resistant infections. RESULTS: Of 100 patients with macrolide-susceptible MG treated with doxycycline + azithromycin, 93 were cured (93.0%; 95% confidence interval [CI], 86.1-97.1). Of 247 patients with macrolide-resistant MG receiving doxycycline + moxifloxacin, 210 were cured (85.0%; 95% CI, 80.0-89.2). parC sequencing was available for 164 (66%) macrolide-resistant infections; 29% had SNPs at parC S83 or D87 (23% S83I). The absence of SNPs at parC S83/D87 was associated with 98.3% cure (95% CI, 93.9-99.8) following doxycycline + moxifloxacin. The presence of the parC S83I-SNP was associated with failure in 62.5% (95% CI, 45.8-77.3). Side effects were common (40%-46%) and predominantly mild and gastrointestinal. CONCLUSIONS: Combination doxycycline + azithromycin achieved high cure for macrolide-susceptible infections. However, in the context of a high prevalence of the parC S83I mutation (23%) in macrolide-resistant infections, doxycycline + moxifloxacin cured only 85%. Infections that were wild-type for S83/D87 experienced high cure following doxycycline + moxifloxacin, supporting the use of a parC-resistance/susceptibility testing strategy in clinical care.

The effect of probiotic supplementation on the gut microbiota of preterm infants.

Introduction. Probiotic supplementation of preterm infants may prevent serious morbidities associated with prematurity.Aim. To investigate the impact of probiotic supplementation on the gut microbiota and determine factors associated with detection of probiotic species in the infant gut.Hypothesis/Gap Statement. Probiotic supplementation increases the long-term colonization of probiotic species in the gut of preterm infants.Methodology. Longitudinal stool samples were collected from a cohort of very preterm infants participating in a blinded randomized controlled trial investigating the impact of probiotic supplementation (containing Bifidobacterium longum subsp. infantis BB-02, Bifidobacterium animalis subsp. lactis BB-12 and Streptococcus thermophilus TH-4) for prevention of late-onset sepsis. The presence of B. longum subsp. infantis, B. animalis subsp. lactis and S. thermophilus was determined for up to 23 months after supplementation ended using real-time PCR. Logistic regression was used to investigate the impact of probiotic supplementation on the presence of each species.Results. Detection of B. longum subsp. infantis [odds ratio (OR): 53.1; 95 % confidence interval (CI): 35.6-79.1; P < 0.001], B. animalis subsp. lactis (OR: 89.1; 95 % CI: 59.0-134.5; P < 0.001) and S. thermophilus (OR: 5.66; 95 % CI: 4.35-7.37; P < 0.001) was increased during the supplementation period in infants receiving probiotic supplementation. Post-supplementation, probiotic-supplemented infants had increased detection of B. longum subsp. infantis (OR: 2.53; 95 % CI: 1.64-3.90; P < 0.001) and B. animalis subsp. lactis (OR: 1.59; 95 % CI: 1.05-2.41; P=0.030). Commencing probiotic supplementation before 5 days from birth was associated with increased detection of the probiotic species over the study period (B. longum subsp. infantis, OR: 1.20; B. animalis subsp. lactis, OR: 1.28; S. thermophilus, OR: 1.45).Conclusion. Probiotic supplementation with B. longum subsp. infantis BB-02, B. animalis subsp. lactis BB-12 and S. thermophilus TH-4 enhances the presence of probiotic species in the gut microbiota of very preterm infants during and after supplementation. Commencing probiotic supplementation shortly after birth may be important for improving the long-term colonization of probiotic species.

Gut microbiota of preterm infants supplemented with probiotics: sub-study of the ProPrems trial.

BACKGROUND: The ProPrems trial, a multi-center, double-blind, placebo-controlled randomized trial, previously reported a 54% reduction in necrotizing enterocolitis (NEC) of Bell stage 2 or more from 4.4 to 2.0% in 1099 infants born before 32 completed weeks' gestation and weighing < 1500 g, receiving probiotic supplementation (with Bifidobacterium longum subsp. infantis BB-02, Streptococcus thermophilus TH-4 and Bifidobacterium animalis subsp. lactis BB-12). This sub-study investigated the effect of probiotic supplementation on the gut microbiota in a cohort of very preterm infants in ProPrems. RESULTS: Bifidobacterium was found in higher abundance in infants who received the probiotics (AOR 17.22; 95% CI, 3.49-84.99, p < 0.001) as compared to the placebo group, and Enterococcus was reduced in infants receiving the probiotic during the supplementation period (AOR 0.27; 95% CI, 0.09-0.82, p = 0.02). CONCLUSION: Probiotic supplementation with BB-02, TH-4 and BB-12 from soon after birth increased the abundance of Bifidobacterium in the gut microbiota of very preterm infants. Increased abundance of Bifidobacterium soon after birth may be associated with reducing the risk of NEC in very preterm infants.