Predictive value for axillary lymph node metastases in early breast cancer: Based on contrast-enhanced ultrasound characteristics of the primary lesion and sentinel lymph node.

OBJECTIVE: To evaluate the value of contrast-enhanced ultrasound (CEUS) characteristics based on primary lesion combined with lymphatic contrast-enhanced ultrasound (LCEUS) patterns of SLN in predicting axillary lymph node metastasis (ALNM) with T1-2N0 breast cancer. METHODS: A retrospective study was conducted in 118 patients with clinically confirmed T1-2N0 breast cancer. Conventional ultrasound (CUS) and CEUS characteristics of the primary lesion and enhancement patterns of SLN were recorded. The risk factors associated with ALNM were selected by univariate and binary logistic regression analysis, and the receiver operating characteristic (ROC) curve was drawn for the evaluation of predictive ALNM metastasis performance. RESULTS: Univariate analysis showed that age, HER-2 status, tumor size, nutrient vessels, extended range of enhancement lesion, and the enhancement patterns of SLN were significant predictive features of ALNM. Further binary logistic regression analysis indicated that the extended range of enhancement lesion (p <  0.001) and the enhancement patterns of SLN (p <  0.001) were independent risk factors for ALNM. ROC analysis showed that the AUC of the combination of these two indicators for predicting ALNM was 0.931 (95% CI: 0.887-0.976, sensitivity: 75.0%, specificity: 99.8%). CONCLUSION: The CEUS characteristics of primary lesion combined with enhancement patterns of SLN are highly valuable in predicting ALNM and can guide clinical axillary surgery decision-making in early breast cancer.

The Value of Percutaneous Contrast-Enhanced Ultrasound in Sentinel Lymph Node Identification, Metastatic Status and Burden Diagnosis in Early Breast Cancer.

OBJECTIVE: The aim of this study was to evaluate the value of percutaneous contrast-enhanced ultrasound (PCEUS) in the identification and characterization of sentinel lymph node (SLN). METHODS: A total of 102 breast cancer patients were collected and underwent preoperative PCEUS, which was used to identify SLN and lymphatic drainage. SLNs were classified into 4 enhancement patterns, including 6 subtypes: homogeneous (I), featured inhomogeneous (II) including inhomogeneous hypoenhancement (IIa) and annular or semi-annular enhancement (IIb), focal filling defect (III) including filling defect area < 50% (IIIa) and filling defect area ≥ 50% (IIIb), and no enhancement (IV). The enhancement patterns of SLNs were compared with the final pathological diagnosis. RESULTS: The identification rate of SLNs using PCEUS was 100% (102/102); the rate of identification of LCs was 100% (102/102), and the coincidence rate was 98.0% (100/102). Four lymphatic drainage patterns (LDPs) including 5 subtypes were found: single LC/single SLN(74.5%), multiple LCs/ single SLN (13.7%) including 2 subtypes:2 LCs/1 SLN and 3 LCs/1 SLN, single LC/multiple SLNs (7.8%), and multiple LCs/multiple SLNs (3.9%). A total of 86.3% (44/51) of patients without axillary metastasis could be safely selected for types I, IIa, and IIb, while the axillary metastasis rates of types III and IV were 74.4% and 87.5%, respectively (P < .001). Compared with grayscale US, the PCEUS significant improvement in diagnosing metastatic SLNs (.794 versus .579, P < .001). For the SLN metastatic burden, Types I, IIa, IIb, and IIIa had ≤2 SLNs metastases, with a pathological coincidence rate of (64/67, 95.5%), and types IIIb and IV had >2 SLNs metastases, with a pathological coincidence rate of (25/35, 71.4%) (P < .001). The AUC of PCEUS for the diagnosis of SLN metastatic status and burden was .794 and .879, respectively (P < .001). CONCLUSION: PCEUS has a high identification rate for SLN and has good potential for diagnosing SLN metastatic status and burden by enhancement patterns.

Contrast-enhanced ultrasound demonstrates greater adjuvant potential: past, current status, and future applications in Hepatocellular carcinoma early diagnosis.

Hepatocellular carcinoma (HCC) has an atypical onset of clinical symptoms and a rapid tumor progression. The majority of patients with HCC are already in the late stages of the disease when they are diagnosed, limiting them to the best available treatments. Contrast-enhanced ultrasound (CEUS) has achieved significant advances in the diagnosis of HCC, including the detection of small lesions, the investigation of more beneficial contrast agents, and the use of CEUS-based radiomics. The purpose of this review is to discuss the relevant research and future challenges of CEUS in the early detection of HCC, to advise more accurate therapy.

Variants of a putative baseplate wedge protein extend the host range of Pseudomonas phage K8.

BACKGROUND: Narrow host range is a major limitation for phage applications, but phages can evolve expanded host range through adaptations in the receptor-binding proteins. RESULTS: Here, we report that Pseudomonas phage K8 can evolve broader host range and higher killing efficiency at the cost of virion stability. Phage K8 host range mutant K8-T239A carries a mutant version of the putative baseplate wedge protein GP075, termed GP075m. While phage K8 adsorbs to hosts via the O-specific antigen of bacterial LPS, phage K8-T239A uses GP075m to also bind the bacterial core oligosaccharide, enabling infection of bacterial strains resistant to K8 infection due to modified O-specific antigens. This mutation in GP075 also alters inter-protein interactions among phage proteins, and reduces the stability of phage particles to environmental stressors like heat, acidity, and alkalinity. We find that a variety of mutations in gp075 are widespread in K8 populations, and that the gp075-like genes are widely distributed among the domains of life. CONCLUSION: Our data show that a typical life history tradeoff occurs between the stability and the host range in the evolution of phage K8. Reservoirs of viral gene variants may be widely present in phage communities, allowing phages to rapidly adapt to any emerging environmental stressors. Video Abstract.

Ultrasound contrast-enhanced patterns of sentinel lymph nodes: predictive value for nodal status and metastatic burden in early breast cancer.

Background: In the post-Z0011 era, sentinel lymph node (SLN) status and metastatic burden determine whether axillary management entails conservative sentinel lymph node biopsy (SLNB) or radical axillary lymph node dissection (ALND) in breast cancer patients. However, SLN status and metastatic burden cannot be evaluated preoperatively in clinical practice. This study explored the predictive value of contrast-enhanced ultrasound (CEUS) patterns of SLN to assess the nodal status and metastatic burden in early breast cancer patients. Methods: A retrospective study was conducted on 88 consecutive patients who were diagnosed with clinical T1-2N0 breast cancer between December 2020 and November 2021 at the Lanzhou University Second Hospital and scheduled for SLNB. Preoperative CEUS was performed to confirm the location and enhancement pattern of the SLN, and the conventional ultrasonic characteristics of the primary breast lesions and SLN were recorded. Intraoperative localized SLN and postoperative pathological results were used as the gold standard for comparison with preoperative ultrasound findings. Results: CEUS successfully identified at least 1 SLN in 88 patients, with a total of 118 SLNs identified in the entire cohort. Univariate analysis showed that lesion size, blood flow grade, SLN longitudinal diameter, cortical thickness, and enhancement pattern were significant predictive features of SLN metastasis. Further multiple regression analysis indicated that the enhancement pattern of the SLN was an independent risk factor for SLN metastasis, with a sensitivity and a specificity of 84.2% (32/38) and 80.0% (40/50), respectively. Meanwhile, the SLN enhancement pattern could predict the lymph node metastasis burden (P<0.001). In patients presenting with a type I (homogeneous enhancement) or type II (heterogeneous enhancement) SLN, 91.5% (65/71) had ≤2 positive SLNs, whereas in patients with a type III (no enhancement) SLN, 70.6% (12/17) had >2 metastatic nodes. Conclusions: The contrast-enhanced pattern of the SLN is an independent risk factor for SLN status. Patients presenting with a type I or type II SLN enhanced pattern are unlikely to have high-burden metastases detected at their final surgical treatment and omission of ALND may be appropriate.

MSMFN: An Ultrasound Based Multi-Step Modality Fusion Network for Identifying the Histologic Subtypes of Metastatic Cervical Lymphadenopathy.

Identifying squamous cell carcinoma and adenocarcinoma subtypes of metastatic cervical lymphadenopathy (CLA) is critical for localizing the primary lesion and initiating timely therapy. B-mode ultrasound (BUS), color Doppler flow imaging (CDFI), ultrasound elastography (UE) and dynamic contrast-enhanced ultrasound provide effective tools for identification but synthesis of modality information is a challenge for clinicians. Therefore, based on deep learning, rationally fusing these modalities with clinical information to personalize the classification of metastatic CLA requires new explorations. In this paper, we propose Multi-step Modality Fusion Network (MSMFN) for multi-modal ultrasound fusion to identify histological subtypes of metastatic CLA. MSMFN can mine the unique features of each modality and fuse them in a hierarchical three-step process. Specifically, first, under the guidance of high-level BUS semantic feature maps, information in CDFI and UE is extracted by modality interaction, and the static imaging feature vector is obtained. Then, a self-supervised feature orthogonalization loss is introduced to help learn modality heterogeneity features while maintaining maximal task-consistent category distinguishability of modalities. Finally, six encoded clinical information are utilized to avoid prediction bias and improve prediction ability further. Our three-fold cross-validation experiments demonstrate that our method surpasses clinicians and other multi-modal fusion methods with an accuracy of 80.06%, a true-positive rate of 81.81%, and a true-negative rate of 80.00%. Our network provides a multi-modal ultrasound fusion framework that considers prior clinical knowledge and modality-specific characteristics. Our code will be available at: https://github.com/RichardSunnyMeng/MSMFN.

Anti-tumor effects of P-LPK-CPT, a peptide-camptothecin conjugate, in colorectal cancer.

To explore highly selective targeting molecules of colorectal cancer (CRC) is a challenge. We previously identified a twelve-amino acid peptide (LPKTVSSDMSLN, namely P-LPK) by phage display technique which may specifically binds to CRC cells. Here we show that P-LPK selectively bind to a panel of human CRC cell lines and CRC tissues. In vivo, Gallium-68 (68Ga) labeled P-LPK exhibits selective accumulation at tumor sites. Then, we designed a peptide-conjugated drug comprising P-LPK and camptothecin (CPT) (namely P-LPK-CPT), and found P-LPK-CPT significantly inhibits tumor growth with fewer side effects in vitro and in vivo. Furthermore, through co-immunoprecipitation and molecular docking experiment, the glutamine transporter solute carrier 1 family member 5 (SLC1A5) was identified as the possible target of P-LPK. The binding ability of P-LPK and SLC1A5 is verified by surface plasmon resonance and immunofluorescence. Taken together, P-LPK-CPT is highly effective for CRC and deserves further development as a promising anti-tumor therapeutic for CRC, especially SLC1A5-high expression type.

Risk Factors for Esophageal Squamous Cell Carcinoma in Patients with Head and Neck Squamous Cell Carcinoma.

Background: Esophageal squamous cell carcinoma (ESCC) is a common second primary neoplasia in patients with a history of head and neck squamous cell carcinoma (HNSCC). The aim of this study was to provide further information and novel insights into the risk factors for ESCC in patients with HNSCC. Methods: We retrospectively analyzed 98 HNSCC patients diagnosed from 2007 to 2017, 30 HNSCC patients suffering from ESCC, who had undergone endoscopic examination because of positive imaging examinations or symptoms, and 68 HNSCC patients who had no ESCC occurrence for at least six years post-HNSCC diagnosis. Associated clinicopathological data and lifestyle information of the ESCC group and the without ESCC group were collected, and a case-control study of risk factors was analyzed between the two groups. Results: The majority (83.4%) of the cases with HNSCC esophageal cancers were male patients over 50 years. We established that 93.75% (30/32) of the ESCC occurred within six years after HNSCC diagnosis. HNSCC location, stage, and radiotherapy history had no significant association with the development of ESCC. High Ki67 labeling index (Ki67 LI) (>46) patients tended to be 3.1 times (95% CI = 1.3-7.6) more likely to develop ESCC compared to low Ki67 LI (≤45) patients (P < 0.05). Drinkers with alcohol flushing response were at a 3.3 times higher risk to have ESCC (95% CI = 1.0-10.4) than drinkers without flush response (P < 0.05). Conclusions: HNSCC patients, especially drinkers with an alcohol flushing response, as well as those with high Ki67 LI of HNSCC tissue, were more likely to develop ESCC.

Exosomal miR-19a decreases insulin production by targeting Neurod1 in pancreatic cancer associated diabetes.

BACKGROUND: New onset diabetes mellitus demonstrates a roughly correlation with pancreatic cancer (PaC), which is unique in PaC and was named as PaC-induced DM, but the inner mechanism remains unclear. Exosomes mediate intercellular communication and bearing microRNAs might be direct constituent of effect in target cells. METHODS AND RESULTS: The isolated exosomes from PaC cells were used to treat pancreatic ß cells or the primary mice islets, and the glucose stimulated insulin secretions were measured. We validated the exosomal miR-19a from PaC cells to be an important mediator in the down regulation of insulin secretion by targeting Neurod1, the validated gene involved in insulin secretion, by using the quantitative real-time PCR, western blot, and promoter luciferase activity. The relative insulin, cAMP and Ca2+ expressions were also assayed to verify the inverse correlation between cancerous miR-19a and pancreatic islets Neurod1. CONCLUSIONS: Our study indicated that signal changes between cancer cells and ß cells via exosomes might be important in the pathogenesis of PaC-induced DM and supplemented the pathogenesis of PaC-induced DM and provide a possible access of PaC screening strategy.

Pancreatic cancer-derived exosomal microRNA-19a induces ß-cell dysfunction by targeting ADCY1 and EPAC2.

New-onset diabetes mellitus has a rough correlation with pancreatic cancer (PaC), but the underlying mechanism remains unclear. This study aimed to explore the exosomal microRNAs and their potential role in PaC-induced ß-cell dysfunction. The pancreatic ß cells were treated with isolated exosomes from PaC cell lines, SW1990 and BxPC-3, before measuring the glucose-stimulated insulin secretion (GSIS), validating that SW1990 and BxPC-3 might disrupt GSIS of both ß cell line MIN6 and primary mouse pancreatic islets. The difference in expression profiles between exosomes and exosome-free medium of PaC cell lines was further defined, revealing that miR-19a secreted by PaC cells might be an important signaling molecule in this process. Furthermore, adenylyl cyclase 1 (Adcy1) and exchange protein directly activated by cAMP 2 (Epac2) were verified as the direct targets of exogenous miR-19a, which was involved in insulin secretion. These results indicated that exosomes might be an important mediator in the pathogenesis of PaC-DM, and miR-19a might be the effector molecule. The findings shed light on the pathogenesis of PaC-DM.

YTHDC1-mediated augmentation of miR-30d in repressing pancreatic tumorigenesis via attenuation of RUNX1-induced transcriptional activation of Warburg effect.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal human cancers. It thrives in a malnourished environment; however, little is known about the mechanisms by which PDAC cells actively promote aerobic glycolysis to maintain their metabolic needs. Gene Expression Omnibus (GEO) was used to identify differentially expressed miRNAs. The expression pattern of miR-30d in normal and PDAC tissues was studied by in situ hybridization. The role of miR-30d/RUNX1 in vitro and in vivo was evaluated by CCK8 assay and clonogenic formation as well as transwell experiment, subcutaneous xenograft model and liver metastasis model, respectively. Glucose uptake, ATP and lactate production were tested to study the regulatory effect of miR-30d/RUNX1 on aerobic glycolysis in PDAC cells. Quantitative real-time PCR, western blot, Chip assay, promoter luciferase activity, RIP, MeRIP, and RNA stability assay were used to explore the molecular mechanism of YTHDC1/miR-30d/RUNX1 in PDAC. Here, we discover that miR-30d expression was remarkably decreased in PDAC tissues and associated with good prognosis, contributed to the suppression of tumor growth and metastasis, and attenuation of Warburg effect. Mechanistically, the m6A reader YTHDC1 facilitated the biogenesis of mature miR-30d via m6A-mediated regulation of mRNA stability. Then, miR-30d inhibited aerobic glycolysis through regulating SLC2A1 and HK1 expression by directly targeting the transcription factor RUNX1, which bound to the promoters of the SLC2A1 and HK1 genes. Moreover, miR-30d was clinically inversely correlated with RUNX1, SLC2A1 and HK1, which function as adverse prognosis factors for overall survival in PDAC tissues. Overall, we demonstrated that miR-30d is a functional and clinical tumor-suppressive gene in PDAC. Our findings further uncover that miR-30d is a novel target for YTHDC1 through m6A modification, and miR-30d represses pancreatic tumorigenesis via suppressing aerobic glycolysis.

Diethyldithiocarbamate-copper complex (CuET) inhibits colorectal cancer progression via miR-16-5p and 15b-5p/ALDH1A3/PKM2 axis-mediated aerobic glycolysis pathway.

Exploring novel anticancer drugs to optimize the efficacy may provide a benefit for the treatment of colorectal cancer (CRC). Disulfiram (DSF), as an antialcoholism drug, is metabolized into diethyldithiocarbamate-copper complex (CuET) in vivo, which has been reported to exert the anticancer effects on various tumors in preclinical studies. However, little is known about whether CuET plays an anti-cancer role in CRC. In this study, we found that CuET had a marked effect on suppressing CRC progression both in vitro and in vivo by reducing glucose metabolism. Mechanistically, using RNA-seq analysis, we identified ALDH1A3 as a target gene of CuET, which promoted cell viability and the capacity of clonal formation and inhibited apoptosis in CRC cells. MicroRNA (miR)-16-5p and 15b-5p were shown to synergistically regulate ALDH1A3, which was negatively correlated with both of them and inversely correlated with the survival of CRC patients. Notably, using co-immunoprecipitation followed with mass spectrometry assays, we identified PKM2 as a direct downstream effector of ALDH1A3 that stabilized PKM2 by reducing ubiquitination. Taken together, we disclose that CuET treatment plays an active role in inhibiting CRC progression via miR-16-5p and 15b-5p/ALDH1A3/PKM2 axis-mediated aerobic glycolysis pathway.

Histone methyltransferase WHSC1 inhibits colorectal cancer cell apoptosis via targeting anti-apoptotic BCL2.

WHSC1 is a histone methyltransferase that facilitates histone H3 lysine 36 dimethylation (H3K36me2), which is a permissive mark associated with active transcription. In this study, we revealed how WHSC1 regulates tumorigenesis and chemosensitivity of colorectal cancer (CRC). Our data showed that WHSC1 as well as H3K36me2 were highly expressed in clinical CRC samples, and high WHSC1 expression is associated with poorer prognosis in OS patients. WHSC1 reduction promoted colon cancer cell apoptosis both in vivo and in vitro. We found that B cell lymphoma-2 (BCL2) expression, an anti-apoptotic protein, is markedly decreased in after WHSC1 depletion. Mechanistic characterization indicated that WHSC1 directly binds to the promoter region of BCL2 gene and regulate its H3K36 dimethylation level. What's more, our study indicated that WHSC1 depletion promotes chemosensitivity in CRC cells. Together, our results suggested that WHSC1 and H3K36me2 modification might be optimal therapeutic targets to disrupt CRC progression and WHSC1-targeted therapy might potentially overcome the resistance of chemotherapeutic agents.

Characteristics of esophageal cancer in patients with head and neck squamous cell carcinoma.

BACKGROUND: We investigated the clinicopathological features of esophageal cancer in patients with a history of head and neck squamous cell carcinoma (HNSCC) with the intention of providing information regarding the characteristics of these patients. METHODS: A retrospective study was performed in 32 cases of esophageal cancer with HNSCC who were diagnosed using upper gastrointestinal endoscopy between 2007 to 2017. Synchronous carcinoma (SC) group and metachronous carcinoma (MC) group was established based on whether esophageal cancer was diagnosed within 6 months after HNSCC diagnosis. The clinicopathological features of esophageal cancer and HNSCC, as well as follow-up treatment and survival, were analyzed in esophageal cancer patients in both groups. RESULTS: There were 8 cases of 8 patients (7 males and 1 female) in the SC group and 24 cases of 22 patients (21 males and 1 female) in the MC group. The majority of esophageal cancer of HNSCC were male patients aged 50-69 years. The average interval time between diagnosis of esophageal cancer and HNSCC was 36.0±39.2 months (3.25±2.19 months for the SC group and 46.90±39.73 months for the MC group). Ninety-three-point-seven-five percent (30/32) of the patients had esophageal cancer within 6 years after HNSCC. The proportion of early esophageal cancer and successful surgical treatment in the SC group was significantly higher compared to the MC group (P<0.05). CONCLUSIONS: Detection of esophageal cancer should be prioritized in HNSCC patients.

The p53-inducible CLDN7 regulates colorectal tumorigenesis and has prognostic significance.

Most colorectal cancer (CRC) are characterized by allele loss of the genes located on the short arm of chromosome 17 (17p13.1), including the tumor suppressor p53 gene. Although important, p53 is not the only driver of chromosome 17p loss. In this study, we explored the biological and prognostic significance of genes around p53 on 17p13.1 in CRC. The Cancer Genome Atlas (TCGA) were used to identify differentially expressed genes located between 1000 kb upstream and downstream of p53 gene. The function of CLDN7 was evaluated by both in vitro and in vivo experiments. Quantitative real-time PCR, western blot, and promoter luciferase activity, immunohistochemistry were used to explore the molecular drivers responsible for the development and progression of CRC. The results showed that CLDN7, located between 1000 kb upstream and downstream of p53 gene, were remarkably differentially expressed in tumor and normal tissues. CLDN7 expression also positively associated with p53 level in different stages of the adenoma-carcinoma sequence. Both in vitro and in vivo assays showed that CLDN7 inhibited cell proliferation in p53 wild type CRC cells, but had no effects on p53 mutant CRC cells. Mechanistically, p53 could bind to CLDN7 promoter region and regulate its expression. Clinically, high CLDN7 expression was negatively correlated with tumor size, invasion depth, lymphatic metastasis and AJCC III/IV stage, but was positively associated with favorable prognosis of CRC patients. Collectively, our work uncovers the tumor suppressive function for CLDN7 in a p53-dependent manner, which may mediate colorectal tumorigenesis induced by p53 deletion or mutation.

Cone beam CT-based measurement of the accessory mental foramina in the Chinese Han population.

Anatomical data of accessory mental foramina (AMFs) were investigated in a Chinese Han population using cone beam CT (CBCT). A retrospective analysis was performed on 527 selected sets of CBCT images. The average frequency and diameter of AMFs, the diameter of the ipsilateral mental foramen (MF), and the center distance and relative position between the AMFs and MF were measured and calculated by three professional dentists. Among the 527 patients, AMFs were identified in 36 cases (frequency 6.83%), of which 68.75% of AMFs were larger than 1 mm. The mean diameters of the AMFs and the ipsilateral MF were 1.32±0.61 mm and 3.26±0.90 mm, respectively. The average distance from the AMFs to the alveolar ridge crest (ARC) was 15.05±3.50 mm, and the average distance to the mandibular plane was 15.87±3.64 mm. The positions of the AMFs relative to the MF varied widely. The AMFs were mostly positioned distal-inferior to the ipsilateral MF and under the mandibular second premolars. Nutrient foramina around the MFs were distinguished from AMFs. The reference plane for measuring AMFs was suggested to be the mandibular plane to increase the repeatability and accuracy of the experiment. Standard planes were proposed to determine the relative position between AMFs and the MFs. Based on our results, we propose that for implant surgeries, the safety region of 2 mm above the MFs should be reevaluated. CBCT examination is recommended before the operation to identify important anatomical structures around the MF region and their variations and set the safety distance on an individual basis.

Biodegradation of Phenanthrene and Heavy Metal Removal by Acid-Tolerant Burkholderia fungorum FM-2.

Phenanthrene (PHE) is a common pollutant of acidic and non-acidic environments that is recalcitrant to biodegradation. Herein, Burkholderia fungorum FM-2 (GenBank accession no. KM263605) was isolated from oil-contaminated soil in Xinjiang and characterized morphologically, physiologically, and phylogenetically. Environmental parameters including PHE concentration, pH, temperature, and salinity were optimized, and heavy metal tolerance was investigated. The MIC of strain FM-2 tolerant to Pb(II) and Cd(II) was 50 and 400 mg L-1, respectively, while the MIC of Zn(II) was >1,200 mg L-1. Atypically for a B. fungorum strain, FM-2 utilized PHE (300 mg L-1) as a sole carbon source over a wide pH range (between pH 3 and 9). PHE and heavy metal metabolism were assessed using gas chromatography (GC), inductively coupled plasma optical emission spectroscopy (ICP-OES), scanning electron microscopy-energy dispersive X-ray spectroscopy (SEM-EDS), Fourier-transform infrared (FTIR) spectroscopy and ultraviolet (UV) absorption spectrometry. The effects of heavy metals on the bioremediation of PHE in soil were investigated, and the findings suggest that FM-2 has potential for combined bioremediation of soils co-contaminated with PHE and heavy metals.