Prognostic significance of absolute lymphocyte count in patients with metastatic renal cell carcinoma receiving first-line combination immunotherapies: results from the International Metastatic Renal Cell Carcinoma Database Consortium.

BACKGROUND: Lymphocytes are closely linked to mechanisms of action of immuno-oncology (IO) agents. We aimed to assess the prognostic significance of absolute lymphocyte count (ALC) in patients with metastatic renal cell carcinoma (mRCC). PATIENTS AND METHODS: Using the International mRCC Database Consortium (IMDC), patients receiving first-line IO-based combination therapy were analysed. Baseline patient characteristics, objective response rates (ORRs), time to next treatment (TTNT), and overall survival (OS) were compared. RESULTS: Of 966 patients included, 195 (20%) had lymphopenia at baseline, and they had a lower ORR (37% versus 45%; P < 0.001), shorter TTNT (10.1 months versus 24.3 months; P < 0.001), and shorter OS (30.4 months versus 48.2 months; P < 0.001). Among 125 patients with lymphopenia at baseline, 52 (42%) experienced ALC recovery at 3 months, and they had longer OS (not reached versus 30.4 months; P = 0.012). On multivariable analysis for OS, lymphopenia was an independent adverse prognostic factor (hazard ratio 1.68; P < 0.001). Incorporation of lymphopenia into the IMDC criteria improved OS prediction accuracy (C-index from 0.688 to 0.707). CONCLUSIONS: Lymphopenia was observed in one-fifth of treatment-naive patients with mRCC and may serve as an indicator of unfavourable oncologic outcomes in the contemporary IO era.

Assessment of Autophagy in Leishmania Parasites.

Leishmaniasis is a neglected tropical disease caused by numerous species of Leishmania parasites, including Leishmania major. The parasite is transmitted by several species of sandfly vectors and infects myeloid cells leading to a myriad of inflammatory responses, immune dysregulations, and disease manifestations. Every cell undergoes autophagy, a self-regulated degradative process that permits the cells to recycle damaged or worn-out organelles in order to maintain cellular health and homeostasis. Studies have shown that Leishmania modulates their host cell autophagic machinery and there are indications that the parasite-specific autophagic processes may be valuable for parasite virulence and survival. However, the role of autophagy in Leishmania is inconclusive because of the limited tools available to study the Leishmania-specific autophagic machinery. Here, we describe methods to study and definitively confirm autophagy in Leishmania major. Transmission electron microscopy (TEM) allowed us to visualize Leishmania autophagosomes, especially those containing damaged mitochondrial content, as well as dividing mitochondria with ongoing fusion/fission processes. Flow cytometry enabled us to identify the amount of acridine orange dye accumulating in the acidic vacuolar compartments in Leishmania major by detecting fluorescence in the red laser when autophagic inhibitors or enhancers were included. These methods will advance studies that aim to understand autophagic regulation in Leishmania parasites that could provide insights into developing improved therapeutic targets against leishmaniasis.

Antigen recognition reinforces regulatory T cell mediated Leishmania major persistence.

Cutaneous Leishmania major infection elicits a rapid T cell response that is insufficient to clear residually infected cells, possibly due to the accumulation of regulatory T cells in healed skin. Here, we used Leishmania-specific TCR transgenic mice as a sensitive tool to characterize parasite-specific effector and immunosuppressive responses in vivo using two-photon microscopy. We show that Leishmania-specific Tregs displayed higher suppressive activity compared to polyclonal Tregs, that was mediated through IL-10 and not through disrupting cell-cell contacts or antigen presentation. In vivo expansion of endogenous Leishmania-specific Tregs resulted in disease reactivation that was also IL-10 dependent. Interestingly, lack of Treg expansion that recognized the immunodominant Leishmania peptide PEPCK was sufficient to restore robust effector Th1 responses and resulted in parasite control exclusively in male hosts. Our data suggest a stochastic model of Leishmania major persistence in skin, where cellular factors that control parasite numbers are counterbalanced by Leishmania-specific Tregs that facilitate parasite persistence.

Quantitative Amplicon Sequencing Is Necessary to Identify Differential Taxa and Correlated Taxa Where Population Sizes Differ.

High-throughput, multiplexed-amplicon sequencing has become a core tool for understanding environmental microbiomes. As researchers have widely adopted sequencing, many open-source analysis pipelines have been developed to compare microbiomes using compositional analysis frameworks. However, there is increasing evidence that compositional analyses do not provide the information necessary to accurately interpret many community assembly processes. This is especially true when there are large gradients that drive distinct community assembly processes. Recently, sequencing has been combined with Q-PCR (among other sources of total quantitation) to generate "Quantitative Sequencing" (QSeq) data. QSeq more accurately estimates the true abundance of taxa, is a more reliable basis for inferring correlation, and, ultimately, can be more reliably related to environmental data to infer community assembly processes. In this paper, we use a combination of published data sets, synthesis, and empirical modeling to offer guidance for which contexts QSeq is advantageous. As little as 5% variation in total abundance among experimental groups resulted in more accurate inference by QSeq than compositional methods. Compositional methods for differential abundance and correlation unreliably detected patterns in abundance and covariance when there was greater than 20% variation in total abundance among experimental groups. Whether QSeq performs better for beta diversity analysis depends on the question being asked, and the analytic strategy (e.g., what distance metric is being used); for many questions and methods, QSeq and compositional analysis are equivalent for beta diversity analysis. QSeq is especially useful for taxon-specific analysis; QSeq transformation and analysis should be the default for answering taxon-specific questions of amplicon sequence data. Publicly available bioinformatics pipelines should incorporate support for QSeq transformation and analysis.

Comparative Metabolomics Profiling Reveals Key Metabolites and Associated Pathways Regulating Tuber Dormancy in White Yam (Dioscorea rotundata Poir.).

Yams are economic and medicinal crops with a long growth cycle, spanning between 9-11 months due to their prolonged tuber dormancy. Tuber dormancy has constituted a major constraint in yam production and genetic improvement. In this study, we performed non-targeted comparative metabolomic profiling of tubers of two white yam genotypes, (Obiaoturugo and TDr1100873), to identify metabolites and associated pathways that regulate yam tuber dormancy using gas chromatography-mass spectrometry (GC-MS). Yam tubers were sampled between 42 days after physiological maturity (DAPM) till tuber sprouting. The sampling points include 42-DAPM, 56-DAPM, 87DAPM, 101-DAPM, 115-DAPM, and 143-DAPM. A total of 949 metabolites were annotated, 559 in TDr1100873 and 390 in Obiaoturugo. A total of 39 differentially accumulated metabolites (DAMs) were identified across the studied tuber dormancy stages in the two genotypes. A total of 27 DAMs were conserved between the two genotypes, whereas 5 DAMs were unique in the tubers of TDr1100873 and 7 DAMs were in the tubers of Obiaoturugo. The differentially accumulated metabolites (DAMs) spread across 14 major functional chemical groups. Amines and biogenic polyamines, amino acids and derivatives, alcohols, flavonoids, alkaloids, phenols, esters, coumarins, and phytohormone positively regulated yam tuber dormancy induction and maintenance, whereas fatty acids, lipids, nucleotides, carboxylic acids, sugars, terpenoids, benzoquinones, and benzene derivatives positively regulated dormancy breaking and sprouting in tubers of both yam genotypes. Metabolite set enrichment analysis (MSEA) revealed that 12 metabolisms were significantly enriched during yam tuber dormancy stages. Metabolic pathway topology analysis further revealed that six metabolic pathways (linoleic acid metabolic pathway, phenylalanine metabolic pathway, galactose metabolic pathway, starch and sucrose metabolic pathway, alanine-aspartate-glutamine metabolic pathways, and purine metabolic pathway) exerted significant impact on yam tuber dormancy regulation. This result provides vital insights into molecular mechanisms regulating yam tuber dormancy.

Variability and genetic merits of white Guinea yam landraces in Nigeria.

Introduction: Landraces represent a significant gene pool of African cultivated white Guinea yam diversity. They could, therefore, serve as a potential donor of important traits such as resilience to stresses as well as food quality attributes that may be useful in modern yam breeding. This study assessed the pattern of genetic variability, quantitative trait loci (QTLs), alleles, and genetic merits of landraces, which could be exploited in breeding for more sustainable yam production in Africa. Methods: A total of 86 white Guinea yam landraces representing the popular landraces in Nigeria alongside 16 elite clones were used for this study. The yam landraces were genotyped using 4,819 DArTseq SNP markers and profiled using key productivity and food quality traits. Results and discussion: Genetic population structure through admixture and hierarchical clustering methods revealed the presence of three major genetic groups. Genome-wide association scan identified thirteen SNP markers associated with five key traits, suggesting that landraces constitute a source of valuable genes for productivity and food quality traits. Further dissection of their genetic merits in yam breeding using the Genomic Prediction of Cross Performance (GPCP) allowed identifying several landraces with high crossing merit for multiple traits. Thirteen landraces were identified as potential genitors to develop segregating progenies to improve multiple traits simultaneously for desired gains in yam breeding. Results of this study provide valuable insights into the patterns and the merits of local genetic diversity which can be utilized for identifying desirable genes and alleles of interest in yam breeding for Africa.

Minimally Invasive Surgery in Port Harcourt, Nigeria: Progress So Far.

Background The global practice of minimally invasive surgery (MIS) has progressed from basic to advanced procedures. Consequent to this, almost all surgical procedures can be performed through a minimally invasive technique. This study aims to audit the practice of MIS in healthcare facilities within a city in a developing country in Africa. Methods This is a multicenter, multispecialty, retrospective descriptive study of minimally invasive diagnostic and therapeutic surgeries performed in private and public health care facilities in Port Harcourt, Rivers State, Nigeria, conducted for a duration of 10 years, from January 2010 to December 2019. A proforma was distributed for completion to identified surgeons from the included study centers. Data on MIS, including types of procedures, time trends, frequency, category of surgery, and cost, were collated. Statistical analysis was performed using IBM Statistical Package for the Social Sciences (IBM SPSS version 20.0, New York, USA). Results There were 5845 minimally invasive procedures performed during the study period, out of which only 92 (1.57%) were carried out in government-owned hospitals. Of these, 2570 were gynecologic (44.0%), 1873 were urologic (32.0%), 1300 were general surgeries (22.2%), 142 were pediatric surgeries (2.4%), and 3 (0.05%) were thoracic minimally invasive procedures performed within the 10-year period. The cost of procedures ranged from <$200 USD to >$2000 USD. The hospital stays ranged from <1 day to a maximum of 13 days. Conclusion The practice of MIS has made significant progress but has been primarily driven by the private sector. Subsidizing the cost of MIS procedures in government-owned hospitals is likely to improve patronage and improve the skills of surgeons.

In vitro and in vivo antimalarial activity and chemical profiling of sugarcane leaves.

Saccharum officinarum Linn. (sugarcane, Family-Poaceae) is employed in Ibibio traditional medicine for the treatment of various infections and diseases such as malaria. We This study aims to assess the antiplasmodial effect of the leaf extract and fractions on human malaria parasite (Plasmodium falciparum) in vitro, and rodent malaria parasite (P. berghei) in vivo, and analyse the bioactive components of the active fraction(s). The leaf extract and fractions of S. officinarum were prepared and their growth inhibitory effects tested against the chloroquine resistant P. falciparum strain (Dd2) and P. berghei infection in mice. An acute toxicity of the extract was determined. A combination of gas chromatography and liquid chromatography-mass spectrometry, and nuclear magnetic resonance spectroscopy was applied for metabolites profiling of crude extract and active fractions. The leaf extract and fractions demonstrated moderate activity against P. falciparum with the dichloromethane fraction producing the most potent activity (EC50 = 15.4 µg/mL). The leaf extract (170-510 mg/kg, p.o., LD50 = 1732 mg/kg) and fractions demonstrated significant (p < 0.05-0.001) effect on P. berghei infection in prophylactic  tests as well as in established infection with n-butanol fractions producing the highest effect. An unusual sulphur-containing compound, dilaurylthiodipropionate, fatty acids, phenolic acids, flavonoid and flavonoid glycoside were identified in the active fractions. These results give credence to the use of sugarcane leaves as malarial remedy locally by confirming the in vitro and in vivo antiplasmodial potential of leaf extract/fractions of S. officinarum.

In vivo antihyperglycaemic and antihyperlipidemic activities and chemical constituents of Solanum anomalum.

Solanum anomalum is a plant used ethnomedically for the treatment of diabetes. The study was aimed to validate ethnomedical claims in rat model and identify the likely antidiabetic compounds. Leaf extract (70-210 mg/kg/day) and fractions (140 mg/kg/day) of S. anomalum were evaluated in hyperglycaemic rats induced using alloxan for effects on blood glucose, lipids and pancreas histology. Phytochemical characterisation of isolated compounds and their identification were performed using mass spectrometry and NMR spectroscopy. Bioinformatics tool was used to predict the possible protein targets of the identified bioactive compounds. The leaf extract/fractions on administration to diabetic rats caused significant lowering of fasting blood glucose of the diabetic rats during single dose study and on repeated administration of the extract. The hydroethanolic leaf extracts also enhanced glucose utilization capacity of the diabetic rats and caused significant lowering of glycosylated hemoglobin levels and elevation of insulin levels in the serum. Furthermore, triglycerides, LDL-cholesterol, and VLDL-cholesterol levels were lowered significantly, while HDL-cholesterol levels were also elevated in the treated diabetic rats. There was absence or few pathological signs in the treated hyperglycaemic rat pancreas compared to that present in the pancreas of control group. Diosgenin, 25(R)-diosgenin-3-O-α-L-rhamnopyranosyl-(1→4)-ß-D-glucopyranoside, uracil, thymine, 1-octacosanol, and octacosane were isolated and identified. Protein phosphatases along with secreted proteins are predicted to be the major targets of diosgenin and the diosgenin glycoside. These results suggest that the leaf extract/fractions of S. anomalum possess antidiabetic and antihyperlipidemic properties, offer protection to the pancreas and stimulate insulin secretion, which can be attributable to the activities of its phytochemical constituents.

Factors Influencing the Risk of Major Amputation in Patients with Diabetic Foot Ulcers Treated by Autologous Cell Therapy.

Introduction: Autologous cell therapy (ACT) is one of the last options for limb salvage in patients with chronic limb-threatening ischemia (CLTI) and diabetic foot ulcers (DFU). However, some patients may still undergo a major amputation even after ACT, but the risk factors for this are not known. Therefore, the aim of our study was to assess the risk factors for major amputation in patients with CLTI and DFU during a 2-year follow-up after ACT. Methods: One hundred and thirteen patients after ACT were included in our study and divided into two groups: Group 1 with major amputation (AMP; n = 37) and Group 2 without amputation (nAMP, n = 76). The risk factors for major amputation were evaluated before ACT and included factors relating to the patient, the DFU, and the cell product. Results: The AMP group had significantly higher C-reactive protein (CRP) levels compared to the nAMP group (22.7 vs. 10.7 mg/L, p = 0.024). In stepwise logistic regression, independent predictors for major amputation were mutation of the gene for methylenetetrahydrofolate reductase (MTHFR) with heterozygote and homozygote polymorphism 1298 (OR 4.33 [95% CI 1.05-17.6]), smoking (OR 3.83 [95% CI 1.18-12.5]), and CRP > 10 mg/L (OR 2.76 [95% CI 0.93-8.21]). Lower transcutaneous oxygen pressure (TcPO2) values were observed in AMP patients compared to the nAMP group at one month (24.5 vs. 33.2, p = 0.012) and at 3 months (31.1 vs. 40.9, p = 0.009) after ACT. Conclusion: Our study showed that the risk for major amputation after ACT in patients with CLTI and DFU is increased by the presence of MTHFR heterozygote and homozygote gene mutations, smoking, and higher CRP at baseline. Lower TcPO2 at one and 3 months after ACT may also have a predictive value. Therefore, it is necessary to stop smoking before ACT, treat any infection, and, above all, consider antiaggregation or anticoagulant treatment after the procedure.

Chromosome evolution and the genetic basis of agronomically important traits in greater yam.

The nutrient-rich tubers of the greater yam, Dioscorea alata L., provide food and income security for millions of people around the world. Despite its global importance, however, greater yam remains an orphan crop. Here, we address this resource gap by presenting a highly contiguous chromosome-scale genome assembly of D. alata combined with a dense genetic map derived from African breeding populations. The genome sequence reveals an ancient allotetraploidization in the Dioscorea lineage, followed by extensive genome-wide reorganization. Using the genomic tools, we find quantitative trait loci for resistance to anthracnose, a damaging fungal pathogen of yam, and several tuber quality traits. Genomic analysis of breeding lines reveals both extensive inbreeding as well as regions of extensive heterozygosity that may represent interspecific introgression during domestication. These tools and insights will enable yam breeders to unlock the potential of this staple crop and take full advantage of its adaptability to varied environments.

PlexinD1 Deficiency in Lung Interstitial Macrophages Exacerbates House Dust Mite-Induced Allergic Asthma.

Interstitial macrophages (IMs) are key regulators of allergic inflammation. We previously showed that the absence of semaphorin 3E (Sema3E) exacerbates asthma features in both acute and chronic asthma models. However, it has not been studied whether Sema3E, via its receptor plexinD1, regulates IM function in allergic asthma. Therefore, we investigated the role of plexinD1 deficiency on IMs in allergic asthma. We found that the absence of plexinD1 in IMs increased airway hyperresponsiveness, airway leukocyte numbers, allergen-specific IgE, goblet cell hyperplasia, and Th2/Th17 cytokine response in the house dust mite (HDM)-induced allergic asthma model. Muc5ac, Muc5b, and α-SMA genes were increased in mice with Plxnd1-deficient IMs compared with wild-type mice. Furthermore, plexinD1-deficient bone marrow-derived macrophages displayed reduced IL-10 mRNA expression, at both the baseline and following HDM challenge, compared with their wild-type counterpart mice. Our data suggest that Sema3E/plexinD1 signaling in IMs is a critical pathway that modulates airway inflammation, airway resistance, and tissue remodeling in the HDM murine model of allergic asthma. Reduced IL-10 expression by plexinD1-deficient macrophages may account for these enhanced allergic asthma features.

Prohibitin plays a role in the functional plasticity of macrophages.

Immunometabolism plays a crucial role in the activation and functional plasticity of immune cells, which in large determines a variety of health and disease states. Factors that integrate immunometabolism in immune cell signaling and functions are beginning to be identified. Previously, we have reported that two transgenic mouse models, Mito-Ob and mutant Mito-Ob (m-Mito-Ob), overexpressing a pleiotropic protein, prohibitin (PHB) or a mutant form of PHB (Tyr114Phe-PHB or m-PHB), respectively, developed distinct immunometabolic phenotypes. Specifically, the immune phenotype appears to be driven by the monocytic cell lineage. Based on immunophenotyping of their splenocytes, we focused our attention on macrophages and hypothesized that PHB may play a role in regulating the two functionally polarized states, M1 and M2. Here, we report that macrophage polarization to the M1 and M2 phenotypes did not alter PHB protein level, but overexpression of PHB in macrophages differentially affected cytokine production in the two polarized states. Furthermore, we found that mutation of the Tyr114 phosphorylation site in PHB affects ERK and STAT6 signaling, arginase synthesis and activity, and mitochondrial respiration in macrophages indicating an important role of PHB in integrating cell signaling events with cell metabolism. In summary, we have discovered that PHB is a crucial regulator in the functional plasticity of macrophages. These initial studies expect to lay the foundation for future research into the relationship between cell signaling events pertaining to immunometabolism in immune cell functions, which are integral components of immune-related health and disease.

Severity of intrauterine adhesions and pregnancy success rates after treatment: Comparison of adhesions obtained from open myomectomy versus uterine curettage.

Intrauterine adhesions (IUA) are rare. A retrospective comparative study was conducted between January 1, 2015, and December 31, 2018. Group A comprised 117 women who developed IUAs after open myomectomy, while Group B comprised 113 women who developed IUAs following uterine trauma caused by uterine instrumentation after a termination of pregnancy (TOP) or spontaneous miscarriage. The IUA grade and pregnancy rates and outcomes were compared using the March classification system. All patients underwent hysteroscopic adhesiolysis. The adhesions tended to be more severe (45/117, 38.5%) in Group A than in Group B (29/113, 25.7%); however, this difference was not statistically significant (Chi-Suare 5.047; p = .080). The period of observation was 24 months from the last hysteroscopy. The pregnancy rate in Group A (26, 22.2%) was significantly lower than in Group B (46, 40.7%) (OR: 2.403, 95% CI: 1.352-4.271; p = .003). Open myomectomy was the preceding aetiological factor in a greater proportion of women with IUA in our study. In cases where pregnancy is desired after open myomectomy, especially where the endometrial cavity is breached, postoperative hysteroscopy to exclude IUAs is recommended.

Severity of intrauterine adhesions and pregnancy success rates after treatment: Comparison of adhesions obtained from open myomectomy versus uterine curettage

Intrauterine adhesions (IUA) are rare. A retrospective comparative study was conducted between January 1, 2015, and December 31, 2018. Group A comprised 117 women who developed IUAs after open myomectomy, while Group B comprised 113 women who developed IUAs following uterine trauma caused by uterine instrumentation after a termination of pregnancy (TOP) or spontaneous miscarriage. The IUA grade and pregnancy rates and outcomes were compared using the March classification system. All patients underwent hysteroscopic adhesiolysis. The adhesions tended to be more severe (45/117, 38.5%) in Group A than in Group B (29/113, 25.7%); however, this difference was not statistically significant (Chi-Suare 5.047; p = .080). The period of observation was 24 months from the last hysteroscopy. The pregnancy rate in Group A (26, 22.2%) was significantly lower than in Group B (46, 40.7%) (OR: 2.403, 95% CI: 1.352­4.271; p = .003). Open myomectomy was the preceding aetiological factor in a greater proportion of women with IUA in our study. In cases where pregnancy is desired after open myomectomy, especially where the endometrial cavity is breached, postoperative hysteroscopy to exclude IUAs is recommended.

Protective CD4+ Th1 cell-mediated immunity is reliant upon execution of effector function prior to the establishment of the pathogen niche.

Intracellular infection with the parasite Leishmania major features a state of concomitant immunity in which CD4+ T helper 1 (Th1) cell-mediated immunity against reinfection coincides with a chronic but sub-clinical primary infection. In this setting, the rapidity of the Th1 response at a secondary site of challenge in the skin represents the best correlate of parasite elimination and has been associated with a reversal in Leishmania-mediated modulation of monocytic host cells. Remarkably, the degree to which Th1 cells are absolutely reliant upon the time at which they interact with infected monocytes to mediate their protective effect has not been defined. In the present work, we report that CXCR3-dependent recruitment of Ly6C+ Th1 effector (Th1EFF) cells is indispensable for concomitant immunity and acute (<4 days post-infection) Th1EFF cell-phagocyte interactions are critical to prevent the establishment of a permissive pathogen niche, as evidenced by altered recruitment, gene expression and functional capacity of innate and adaptive immune cells at the site of secondary challenge. Surprisingly, provision of Th1EFF cells after establishment of the pathogen niche, even when Th1 cells were provided in large quantities, abrogated protection, Th1EFF cell accumulation and IFN-γ production, and iNOS production by inflammatory monocytes. These findings indicate that protective Th1 immunity is critically dependent on activation of permissive phagocytic host cells by preactivated Th1EFF cells at the time of infection.

Visualizing the In Vivo Dynamics of Anti-Leishmania Immunity: Discoveries and Challenges.

Intravital microscopy, such as 2-photon microscopy, is now a mainstay in immunological research to visually characterize immune cell dynamics during homeostasis and pathogen infections. This approach has been especially beneficial in describing the complex process of host immune responses to parasitic infections in vivo, such as Leishmania. Human-parasite co-evolution has endowed parasites with multiple strategies to subvert host immunity in order to establish chronic infections and ensure human-to-human transmission. While much focus has been placed on viral and bacterial infections, intravital microscopy studies during parasitic infections have been comparatively sparse. In this review, we will discuss how in vivo microscopy has provided important insights into the generation of innate and adaptive immunity in various organs during parasitic infections, with a primary focus on Leishmania. We highlight how microscopy-based approaches may be key to providing mechanistic insights into Leishmania persistence in vivo and to devise strategies for better parasite control.

Cultivation and Genome Sequencing of Bacteria Isolated From the Coffee Berry Borer (Hypothenemus hampei), With Emphasis on the Role of Caffeine Degradation.

The coffee berry borer, the most economically important insect pest of coffee worldwide, is the only insect capable of feeding and reproducing solely on the coffee seed, a food source containing the purine alkaloid caffeine. Twenty-one bacterial species associated with coffee berry borers from Hawai'i, Mexico, or a laboratory colony in Maryland (Acinetobacter sp. S40, S54, S55, Bacillus aryabhattai, Delftia lacustris, Erwinia sp. S38, S43, S63, Klebsiella oxytoca, Ochrobactrum sp. S45, S46, Pantoea sp. S61, Pseudomonas aeruginosa, P. parafulva, and Pseudomonas sp. S30, S31, S32, S37, S44, S60, S75) were found to have at least one of five caffeine N-demethylation genes (ndmA, ndmB, ndmC, ndmD, ndmE), with Pseudomonas spp. S31, S32, S37, S60 and P. parafulva having the full complement of these genes. Some of the bacteria carrying the ndm genes were detected in eggs, suggesting possible vertical transmission, while presence of caffeine-degrading bacteria in frass, e.g., P. parafulva (ndmABCDE) and Bacillus aryabhattai (ndmA) could result in horizontal transmission to all insect life stages. Thirty-five bacterial species associated with the insect (Acinetobacter sp. S40, S54, S55, B. aryabhattai, B. cereus group, Bacillus sp. S29, S70, S71, S72, S73, D. lacustris, Erwinia sp. S38, S43, S59, S63, K. oxytoca, Kosakonia cowanii, Ochrobactrum sp. S45, S46, Paenibacillus sp. S28, Pantoea sp. S61, S62, P. aeruginosa, P. parafulva, Pseudomonas sp. S30, S31, S32, S37, S44, S60, S75, Stenotrophomonas sp. S39, S41, S48, S49) might contribute to caffeine breakdown using the C-8 oxidation pathway, based on presence of genes required for this pathway. It is possible that caffeine-degrading bacteria associated with the coffee berry borer originated as epiphytes and endophytes in the coffee plant microbiota.