Bioremediation of lead and chromium from wastewater by Pseudomonas putida and Bacillus subtilis isolated from oil contaminated soil: A comparative study.

Research Article | DOI: https://doi.org/10.31579/2766-2314/170

Bioremediation of lead and chromium from wastewater by Pseudomonas putida and Bacillus subtilis isolated from oil contaminated soil: A comparative study.

  • Safaa 1*
  • Ali Emad 1
  • Abdalameer 2

Institute of Genetic Engineering and Biotechnology for postgraduate studies, University of Baghdad, Baghdad, Iraq 

*Corresponding Author: Safaa., Institute of Genetic Engineering and Biotechnology for postgraduate studies, University of Baghdad, Baghdad, Iraq

Citation: Safaa, Abdalameer, Ali Emad (2025), Research Article: Exploring the Relationship Between Family Income and Youth Substance Abuse., J, Biotechnology and Bioprocessing, 6(5): DOI: 10.31579/2766-2314/170

Copyright: © 2025 Safaa, this is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Received: 01 October 2025 | Accepted: 08 October 2025 | Published: 15 October 2025

Keywords:

Abstract

This study investigated the bioremediation potential of Pseudomonas putida and Bacillus subtilis, isolated from oil-contaminated soil, for removing lead and chromium from synthetic wastewater. The bacteria were identified using biochemical tests and 16S rDNA sequencing. Maximum tolerance levels to chromium and lead were determined for multiple isolates of each species. Bioremediation experiments were conducted over 8 days using pure cultures of each bacterium and a mixed culture. P. putida showed higher tolerance and removal efficiency for both metals compared to B. subtilis. The mixed culture demonstrated synergistic effects, achieving 93.6% chromium removal and 91.6% lead removal after 8 days, outperforming the individual cultures. This study highlights the potential of using P. putida and B. subtilis, particularly in combination, for effective bioremediation of chromium and lead from contaminated wastewater. Further optimization of growth conditions and metabolic pathways could enhance the heavy metal removal efficiency of these bacterial species.

Introduction

Heavy metals such as lead and chromium are common contaminants in industrial and municipal wastewaters, posing serious threats to the environment and public health due to their toxic nature, non-biodegradability, and accumulative behaviors (Al-Enezi, Hamoda, and Fawzi 2004). These metals can enter wastewater through industrial discharges, household chemicals, and various anthropogenic activities like metal finishing, paint manufacturing, and electroplating (Dongre 2021).

Interestingly, while chromium in its trivalent form (Cr3+) is vital for metabolism in small quantities, its hexavalent form (Cr6+) is highly toxic, carcinogenic, and mutagenic (Laxmi and Kaushik 2020). Similarly, lead, even in small amounts, can cause acute health disorders when ingested beyond stringent levels (Feng et al. 2018). Both metals can bioaccumulate in the food chain, ultimately affecting human health through contaminated water and food sources (Sharifuzzaman et al. 2016). Various methods have been developed for the removal of heavy metals from wastewater, including ion exchange, membrane filtration, chemical precipitation, and adsorption (Muchie and Akpor 2010). 

Biological approaches, such as bioremediation, are emerging as eco-friendly alternatives to physicochemical metho (Laxmi and Kaushik 2020). Bioremediation using microorganisms has emerged as a promising, cost-effective, and environmentally friendly approach for removing heavy metals from contaminated environments. Microbes possess various mechanisms for metal sequestration, enabling them to have high metal biosorption capacities (Kulshreshtha et al. 2014). This method is considered safer, cleaner, and more effective than traditional physicochemical approaches for decontaminating sites polluted with a wide range of contaminants (Ayangbenro and Babalola 2017). Bioremediation presents a viable alternative to conventional heavy metal removal techniques, offering a sustainable solution to environmental pollution. As research in this field continues to evolve, bioremediation is poised to play a crucial role in environmental protection and sustainable development (Fouda-Mbanga et al. 2024).

Pseudomonas putida is a highly versatile bacterium with remarkable metabolic capabilities, enabling it to adapt to various environmental niches and effectively degrade a wide range of pollutants, including heavy metals. The genome sequences of several P. putida strains have revealed their diverse metabolic activities and ability to thrive in contaminated environments (Wu et al. 2011). 

Chromate reductase enzymes play a crucial role in bacterial resistance to hexavalent chromium (Cr (VI)) by facilitating its reduction to the less toxic trivalent chromium (Cr (III)). These enzymes are widely distributed among various bacterial species and are considered a key strategy for detoxifying Cr(VI) (Ghosh, Jasu, and Ray 2021).Interestingly, the efficiency of chromate reduction varies among different bacterial strains. For instance, Lactobacillus strains have been reported to completely reduce 32 ppm of Cr(VI) within 6 to 8 hours, making them faster in Cr(VI) reduction than any other known bacteria (Mishra et al. 2012) . 

Similarly, Escherichia coli FACU demonstrated a chromate reductase specific activity of 361.33 μmol/L of Cr(VI) per min per mg protein (Mohamed et al. 2020).

Chromate reductase enzymes are essential for bacterial chromium detoxification and bioremediation. These enzymes not only protect the bacteria from Cr (VI) toxicity but also offer potential applications in environmental cleanup. The development of immobilized enzymatic systems (Robins et al. 2013). Bacillus subtilis has indeed been extensively studied for its ability to sequester heavy metals through biosorption and bioaccumulation. Multiple studies have demonstrated its effectiveness in removing various heavy metals from contaminated environments. Bacillus subtilis has shown significant potential for heavy metal removal through biosorption, which is a cell surface phenomenon [15]. The biosorption process is influenced by factors such as pH, temperature, and contact time. Optimal conditions for B. subtilis include a pH range of 5.0-6.0 and a contact time of 30 minutes (Lorestani, Merrikhpour, and Cheraghi 2020). B. subtilis has also demonstrated the ability to bioaccumulate heavy metals intracellular (Jin et al. 2023).

The effectiveness of Bacillus subtilis in removing heavy metals, coupled with its adaptability to various environmental conditions, highlights its potential as a sustainable solution for addressing heavy metal contamination in different environments. The integration of these bacterial species into bioremediation processes offers a sustainable and eco-friendly alternative to conventional chemical methods. Ongoing research continues to explore the genetic and biochemical pathways involved in metal resistance and detoxification, aiming to optimize bioremediation strategies for industrial applications.

The main goal in this study is to investigate Pseudomonas putida and Bacillus subtilis bioremediation activity for lead and chromium under experiment operation conditions.

Materials and Methods

Samples Collection and bacteria isolation and identification

Soil sample was collected from contaminated site with oil from Refining and Petrochemical Company, Al-Dora, Baghdad, Iraq to isolate bacteria.

Using the spread plate method, bacteria were separated from the oil-contaminated soil and placed on nutrient agar medium. A flask holding nine milliliters of regular saline solution was filled with one gram of soil sample, which was then serially diluted. Each suspension was spread out and plated in 10 μL on solid nutrient agar. Following a 24-hour incubation period at 35°C, the bacterial colonies were isolated and maintained on nutrient agar slants for subsequent investigations.

To identify isolated colonies of heavy metal-tolerant bacteria, characteristics such as their elevation, shape, and superficial forms were noted, also biochemical testing and Gram staining(Singh and Hiranmai 2021).

Extraction of DNA and PCR amplification and sequencing of 16S rDNA

The targeted bacterial DNA was extracted by Macrogen Universal Kit according to the reported procedure (Macrogen Company Korea). The 16S rRNA genes were PCR-amplified from the genomic DNA using the bacterial Macrogen universal primer for Pseudomonas putida and Bacillus subtilis as shown in Table 1.

Pseudomonas putida
Primer typetypeSequence 
518FUniversal Forward5'CCAGCAGCCGCGGTAATACG-3'
800RUniversal Reverse 5' TACCAGGGTATCTAATCC -3'
Bacillus subtilis
Primer typetypeSequence 
27FUniversalForward5′AGAGTTTGATCMTGGCTCAG -3'
1492RUniversal Reverse 5′-TACCTTGTTACGACTT -3'

Table 1: Primers of 16S rDNA used in the study.

Inoculum preparation

For every isolate and the consortium, a standardized bacterial-saline medium suspension was created. The bacterial cells from each isolate were pelleted by centrifugation at 4000 rpm for 15 min at 4oC using an Eppendorf Centrifuge, after each isolate was grown for 24 hours in nutrient broth. After two rinses, the pellet was resuspended with 0.85% NaCl to yield an OD600nm of 0.5. To create the consortium culture, each single isolate was mixed in an equal portion and topped off with 0.85% NaCl. This resulted in an end reading of OD600nm 0.5, which, according to the spread plate method, corresponds to roughly 1*107 CFU per ml. Every experiment that came after it started with this culture (Wong et al. 2015)

Evaluation of bacterial strains for chromium and lead tolerance: 

Using nutrient agar and the agar plate dilution method (Bergey 1994), the resistant to chromium and lead (Cr (VI) and PbCl2) of the isolated bacterial strains was assessed. Spot inoculation (10 μL) of 108 mLG1 cells was performed on freshly prepared agar plates amended with increasing concentrations of Chromium (0-1400 μg/mL) and Lead (0-1000 μg/mL). The Maximum Resistance Level (MRL) was determined by incubating plates at 35°C for 72 hours, during which time the highest concentration of heavy metals that supported growth was found. Three copies of each experiment were conducted.

Chromium and lead reduction by bacterial isolates: 

A 15-minute autoclave sterilization at 121 degrees Celsius was used on the wastewater. 100 ml of the sterilized wastewater was added to each of the seven 120 ml sterile conical flasks. The first flask (Control) was left uninoculated, while the second and third were infused with 20μl of a 24-hour nutrient broth culture of Bacillus subtilis and Pseudomonas putida, respectively. Two different cultures of the bacteria were added to the fourth flask for inoculation. An atomic absorption spectrophotometer was used to measure the concentrations of lead and chromium after the experiment was allowed to stand for 24 hours. When comparisons between the values obtained before and after treatment were made, percentage reductions of heavy metals were found. One milliliter of each flask's culture was centrifuged (6000 rpm) for ten minutes at 10°C in order to reduce lead and chromium. The amount of Cr (VI) in the supernatant was then measured using the 1,5-diphenyl carbazide method (Federation 1999).

For eight days, the flasks were incubated on a New Brunswick gyratory shaker. Samples were taken out every two days so that the atomic absorption spectrophotometer could be used for analysis.

Results and Discussion

Table 2 presents the results of biochemical tests performed on bacterial isolates from oil-contaminated soil. These tests are crucial for identifying and differentiating between Bacillus subtilis and Pseudomonas putida. B. subtilis: Gram-positive (G+ve) bacilli (rod-shaped). This indicates that B. subtilis has a thick peptidoglycan layer in its cell wall, which retains the crystal violet stain during the Gram staining process. Furthermore, P. putida: Gram-negative (G-ve) rods. This confirms that P. putida has a thinner peptidoglycan layer and an outer membrane, which doesn't retain the crystal violet stain.

Both B. subtilis and P. putida isolates are motile. This indicates that both species possess flagella, enabling them to move in their environment, which can be important for accessing nutrients and escaping adverse conditions in the soil. B. subtilis is Positive for spore formation. This means that B. subtilis can form endospores, Nevertheless, P. putida is Negative for spore formation. P. putida does not form endospores. Both B. subtilis and P. putida are catalase-positive. This indicates both species produce the enzyme catalase, which breaks down hydrogen peroxide (a toxic byproduct of aerobic metabolism) into water and oxygen. This allows them to survive in oxygen-rich environments. B. subtilis is Negative for oxidase. This means it lacks the enzyme cytochrome oxidase, which is part of the electron transport chain. P. putida: Positive for oxidase. This indicates the presence of cytochrome oxidase. This is a crucial test for differentiating between these two genera. Pseudomonas are generally oxidase-positive, while Bacillus are oxidase-negative. 

Both B.subtilis and P.putida are positive for gelatin hydrolysis. This indicates that both bacteria produce gelatinase, an enzyme that breaks down gelatin, which provides them with nutrients. B. subtilis was positive for starch hydrolysis. It produces amylase, an enzyme that breaks down starch. But P. putida was Negative for starch hydrolysis Both B. subtilis and P. putida were positive for Simmons Citrate utilization. This indicates that both bacteria can use citrate as their sole carbon source, demonstrating their metabolic versatility. B. subtilis was Negative for fluorescence. It does not produce fluorescent pigments. While, P. putida was fluorescence due to fluorescent pigments generation.

Figure 1 depicts the maximum tolerance levels of Pseudomonas putida to chromium and lead. P. putida showed the highest tolerance to chromium, with maximum tolerance levels ranging from 600 to 1200 μg/ml. Tolerance to lead was significantly lower, ranging from 100 to 400 μg/ml. Tolerance levels varied across the 6 isolates (P1 to P6), with P2 and P3 exhibiting the highest chromium tolerance at 1200 and 1000 μg/ml respectively. Lead tolerance was relatively consistent across isolates, staying below 400 μg/ml. This data indicates that the Pseudomonas putida isolates tested have a much greater capacity to tolerate chromium compared to lead in their environment. The substantial variability in chromium tolerance between isolates also suggests genetic diversity in chromium resistance mechanisms within this bacterial species.

The maximum tolerance levels of Bacillus subtilis isolates to the heavy metals chromium and lead was measured by the agar plate dilution method can be dispatched in Figure 2. Chromium tolerance ranged from 100 to 1000 μg/ml across the 8 isolates (B1 to B8). Lead tolerance was significantly lower, ranging from 100 to 400 μg/ml. The highest chromium tolerance was seen in isolates B5 and B7, at 1000 μg/ml each. Lead tolerance was relatively consistent across isolates, mostly staying below 300 units. Bacillus subtilis isolates demonstrated a greater capacity to tolerate chromium compared to lead. The substantial variability in chromium tolerance across isolates suggests genetic diversity in chromium resistance mechanisms within this bacterial species.

The data presented in Table 3 shows the reduction of chromium and lead concentrations in a heavy metal synthetic water sample using a pure culture of Bacillus subtilis over an 8-day period. The initial chromium concentration was 1000 μg/ml, which decreased by 5.5% to 945 μg/ml by day 2. The chromium level was further reduced by 29.5% to 705 μg/ml on day 4, 43% to 570 μg/ml on day 6, and 50.8% to 492 μg/ml on day 8. The initial lead concentration was 300 μg/ml, which decreased by 3.3% to 290 μg/ml on day 2, 31.3% to 206 μg/ml on day 4, 45.3% to 164 μg/ml on day 6, and 56.6% to 130 μg/ml on day 8. The Bacillus subtilis culture demonstrated a greater capacity to reduce lead levels compared to chromium over the 8-day period.

Recent finding by Wrobel highlighted that Bacillus subtilis can effectively remove various heavy metals, including chromium and lead, through biosorption and intracellular accumulation.(Wróbel et al. 2023) Additionally, (Tarangini et al. 2009) reported that optimizing growth conditions and metabolic pathways in Bacillus subtilis can further enhance its heavy metal removal efficiency.

Table 3: Reduction of Chromium and Lead in heavy metal synthetic water using a pure culture of Bacillus subtilis. presents the bioremediation of chromium and lead by a Pseudomonas putida culture in a heavy metal synthetic water sample over 8 days. The initial chromium concentration was 1200 μg/ml, decreasing by 20.4% to 955 μg/ml on day 2, 46.4% to 643 μg/ml on day 4, 73.1% to 323 μg/ml on day 6, and 82.5% to 210 μg/ml on day 8. The initial lead concentration was 500 μg/ml, reducing by 36.2% to 319 μg/ml on day 2, 46% to 270 μg/ml on day 4, 73.4% to 133 μg/ml on day 6, and 81% to 95 μg/ml on day 8. The P. putida culture demonstrated greater capacity to remove lead compared to chromium, with lead decreasing over 81% and chromium 82.5%.

it was found P. putida can effectively biosorb and accumulate chromium (Olaya-Abril et al. 2024), while Ruparelia and Patel (2024) reviewed bacterial mechanisms for metal bioremediation, including Pseudomonas. Optimizing growth conditions and metabolic pathways in P. putida can further enhance its heavy metal removal efficiency (Abbasi et al. 2024).

The bioremediation of chromium and lead in a heavy metal synthetic water using a mixed culture of Pseudomonas putida and Bacillus subtilis can be demonstrated in Table 4:Bioremediation of Chromium and Lead in heavy metal synthetic water using a pure culture of Pseudomonas putida.. The initial chromium concentration was 1000 μg/ml, decreasing by 32.6% to 674 μg/ml on day 2, 59.8% to 402 μg/ml on day 4, 84.3% to 157 μg/ml on day 6, and 93.6% to 64 μg/ml on day 8. The initial lead concentration was 300 μg/ml, reducing by 45.3% to 164 μg/ml on day 2, 57.7% to 127 μg/ml on day 4, 74.7% to 76 μg/ml on day 6, and 91.6% to 25 μg/ml on day 8. The mixed culture demonstrated greater capacity to remove chromium compared to lead, with chromium decreasing over 93% and lead 91.6%.

Papers reported that bioremediation potential can be enhanced when of Pseudomonas-Bacillus consortia was applied for heavy metals removal. Kumar (2024) found the synergistic interactions between these bacteria improve metal biosorption and intracellular accumulation. Similarly Alotaibi, Khan, and Shamim (2021) reported that optimizing growth conditions and metabolic pathways in mixed cultures can further enhance heavy metal removal efficiency.

The results of DNA extraction, PCR amplification, and 16S rDNA sequencing for Pseudomonas putida and Bacillus subtilis are as follows: DNA was successfully extracted from the bacterial samples using the Macrogen Universal Kit. PCR amplification of the 16S rRNA genes was performed using specific universal primers for each bacterial species. For Pseudomonas putida, the universal forward primer 518F (5'-CCAGCAGCCGCGGTAATACG-3') and the universal reverse primer 800R (5'-TACCAGGGTATCTAATCC-3') were used. These primers amplified a region of approximately 282 base pairs within the 16S rRNA gene. For Bacillus subtilis, the universal forward primer 27F (5'-AGAGTTTGATCMTGGCTCAG-3') and the universal reverse primer 1492R (5'-TACCTTGTTACGACTT-3') were employed. This primer pair amplified a larger region of approximately 1465 base pairs within the 16S rRNA gene. The PCR products were successfully amplified and visualized using gel electrophoresis, confirming the presence of the expected DNA fragments for both bacterial species. The amplified 16S rDNA sequences were then purified and sent for sequencing to determine the exact nucleotide composition of the amplified regions.

Biochemical testB. subtilisP. putida
Gram StainingG+ve bacilliG-ve rods
Motilitymotilemotile
Spore formation+-
Catalase++
Oxidase-+
Gelatin hydrolysis++
Starch hydrolysis+-
Simmons Citrate++
Fluorescence-+

Table 2: Results of Biochemical tests of bacterial isolated from oil contaminated soil (8 isolates of B. subtilis and 6 isolates of P. putida)

Figure 1: Maximum tolerance level of Pseudomonas putida isolates to chromium and Lead by agar plate dilution method.

Figure 2: Maximum tolerance level of Bacillus subtilis isolates to chromium and Lead by agar plate dilution method.

Heavy metalInitial Concentration (μg/ml)

Day 2

 (reduction %)

Day 4 (reduction %)

Day 6

(reduction %)

Day 8

(reduction %)

Chromium1000 945 (5.5%)705 (29.5%)570 (43%)492 (50.8%)
Lead300290206164130 (56.6%)

Table 3: Reduction of Chromium and Lead in heavy metal synthetic water using a pure culture of Bacillus subtilis.

Heavy metalInitial Concentration (μg/ml)

Day 2

 (reduction %)

Day 4 (reduction %)

Day 6

(reduction %)

Day 8

(reduction %)

Chromium1200 955643323210 (82.5%)
Lead50031927013395 (81%)

Table 4:Bioremediation of Chromium and Lead in heavy metal synthetic water using a pure culture of Pseudomonas putida.

Heavy metalInitial Concentration (μg/ml)

Day 2

 (reduction %)

Day 4 (reduction %)

Day 6

(reduction %)

Day 8

(reduction %)

Chromium1000 67440215764 (93.6%)
Lead3001641277625 (91.6%)

Table 5: Bioremediation of Chromium and Lead in heavy metal synthetic water using a mixed culture of Pseudomonas putida and Bacillus subtilis.

References

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George Varvatsoulias

Dear editorial department: On behalf of our team, I hereby certify the reliability and superiority of the International Journal of Clinical Case Reports and Reviews in the peer review process, editorial support, and journal quality. Firstly, the peer review process of the International Journal of Clinical Case Reports and Reviews is rigorous, fair, transparent, fast, and of high quality. The editorial department invites experts from relevant fields as anonymous reviewers to review all submitted manuscripts. These experts have rich academic backgrounds and experience, and can accurately evaluate the academic quality, originality, and suitability of manuscripts. The editorial department is committed to ensuring the rigor of the peer review process, while also making every effort to ensure a fast review cycle to meet the needs of authors and the academic community. Secondly, the editorial team of the International Journal of Clinical Case Reports and Reviews is composed of a group of senior scholars and professionals with rich experience and professional knowledge in related fields. The editorial department is committed to assisting authors in improving their manuscripts, ensuring their academic accuracy, clarity, and completeness. Editors actively collaborate with authors, providing useful suggestions and feedback to promote the improvement and development of the manuscript. We believe that the support of the editorial department is one of the key factors in ensuring the quality of the journal. Finally, the International Journal of Clinical Case Reports and Reviews is renowned for its high- quality articles and strict academic standards. The editorial department is committed to publishing innovative and academically valuable research results to promote the development and progress of related fields. The International Journal of Clinical Case Reports and Reviews is reasonably priced and ensures excellent service and quality ratio, allowing authors to obtain high-level academic publishing opportunities in an affordable manner. I hereby solemnly declare that the International Journal of Clinical Case Reports and Reviews has a high level of credibility and superiority in terms of peer review process, editorial support, reasonable fees, and journal quality. Sincerely, Rui Tao.

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Rui Tao

Clinical Cardiology and Cardiovascular Interventions I testity the covering of the peer review process, support from the editorial office, and quality of the journal.

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Khurram Arshad

Clinical Cardiology and Cardiovascular Interventions, we deeply appreciate the interest shown in our work and its publication. It has been a true pleasure to collaborate with you. The peer review process, as well as the support provided by the editorial office, have been exceptional, and the quality of the journal is very high, which was a determining factor in our decision to publish with you.

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Gomez Barriga Maria Dolores

The peer reviewers process is quick and effective, the supports from editorial office is excellent, the quality of journal is high. I would like to collabroate with Internatioanl journal of Clinical Case Reports and Reviews journal clinically in the future time.

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Lin Shaw Chin

Clinical Cardiology and Cardiovascular Interventions, I would like to express my sincerest gratitude for the trust placed in our team for the publication in your journal. It has been a true pleasure to collaborate with you on this project. I am pleased to inform you that both the peer review process and the attention from the editorial coordination have been excellent. Your team has worked with dedication and professionalism to ensure that your publication meets the highest standards of quality. We are confident that this collaboration will result in mutual success, and we are eager to see the fruits of this shared effort.

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Maria Dolores Gomez Barriga

Dear Dr. Jessica Magne, Editorial Coordinator 0f Clinical Cardiology and Cardiovascular Interventions, I hope this message finds you well. I want to express my utmost gratitude for your excellent work and for the dedication and speed in the publication process of my article titled "Navigating Innovation: Qualitative Insights on Using Technology for Health Education in Acute Coronary Syndrome Patients." I am very satisfied with the peer review process, the support from the editorial office, and the quality of the journal. I hope we can maintain our scientific relationship in the long term.

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Dr Maria Dolores Gomez Barriga

Dear Monica Gissare, - Editorial Coordinator of Nutrition and Food Processing. ¨My testimony with you is truly professional, with a positive response regarding the follow-up of the article and its review, you took into account my qualities and the importance of the topic¨.

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Dr Maria Regina Penchyna Nieto

Dear Dr. Jessica Magne, Editorial Coordinator 0f Clinical Cardiology and Cardiovascular Interventions, The review process for the article “The Handling of Anti-aggregants and Anticoagulants in the Oncologic Heart Patient Submitted to Surgery” was extremely rigorous and detailed. From the initial submission to the final acceptance, the editorial team at the “Journal of Clinical Cardiology and Cardiovascular Interventions” demonstrated a high level of professionalism and dedication. The reviewers provided constructive and detailed feedback, which was essential for improving the quality of our work. Communication was always clear and efficient, ensuring that all our questions were promptly addressed. The quality of the “Journal of Clinical Cardiology and Cardiovascular Interventions” is undeniable. It is a peer-reviewed, open-access publication dedicated exclusively to disseminating high-quality research in the field of clinical cardiology and cardiovascular interventions. The journal's impact factor is currently under evaluation, and it is indexed in reputable databases, which further reinforces its credibility and relevance in the scientific field. I highly recommend this journal to researchers looking for a reputable platform to publish their studies.

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Dr Marcelo Flavio Gomes Jardim Filho

Dear Editorial Coordinator of the Journal of Nutrition and Food Processing! "I would like to thank the Journal of Nutrition and Food Processing for including and publishing my article. The peer review process was very quick, movement and precise. The Editorial Board has done an extremely conscientious job with much help, valuable comments and advices. I find the journal very valuable from a professional point of view, thank you very much for allowing me to be part of it and I would like to participate in the future!”

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Zsuzsanna Bene

Dealing with The Journal of Neurology and Neurological Surgery was very smooth and comprehensive. The office staff took time to address my needs and the response from editors and the office was prompt and fair. I certainly hope to publish with this journal again.Their professionalism is apparent and more than satisfactory. Susan Weiner

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Dr Susan Weiner

My Testimonial Covering as fellowing: Lin-Show Chin. The peer reviewers process is quick and effective, the supports from editorial office is excellent, the quality of journal is high. I would like to collabroate with Internatioanl journal of Clinical Case Reports and Reviews.

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Lin-Show Chin

My experience publishing in Psychology and Mental Health Care was exceptional. The peer review process was rigorous and constructive, with reviewers providing valuable insights that helped enhance the quality of our work. The editorial team was highly supportive and responsive, making the submission process smooth and efficient. The journal's commitment to high standards and academic rigor makes it a respected platform for quality research. I am grateful for the opportunity to publish in such a reputable journal.

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Sonila Qirko

My experience publishing in International Journal of Clinical Case Reports and Reviews was exceptional. I Come forth to Provide a Testimonial Covering the Peer Review Process and the editorial office for the Professional and Impartial Evaluation of the Manuscript.

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Luiz Sellmann

I would like to offer my testimony in the support. I have received through the peer review process and support the editorial office where they are to support young authors like me, encourage them to publish their work in your esteemed journals, and globalize and share knowledge globally. I really appreciate your journal, peer review, and editorial office.

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Zhao Jia

Dear Agrippa Hilda- Editorial Coordinator of Journal of Neuroscience and Neurological Surgery, "The peer review process was very quick and of high quality, which can also be seen in the articles in the journal. The collaboration with the editorial office was very good."

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Thomas Urban

I would like to express my sincere gratitude for the support and efficiency provided by the editorial office throughout the publication process of my article, “Delayed Vulvar Metastases from Rectal Carcinoma: A Case Report.” I greatly appreciate the assistance and guidance I received from your team, which made the entire process smooth and efficient. The peer review process was thorough and constructive, contributing to the overall quality of the final article. I am very grateful for the high level of professionalism and commitment shown by the editorial staff, and I look forward to maintaining a long-term collaboration with the International Journal of Clinical Case Reports and Reviews.

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Cristina Berriozabal

To Dear Erin Aust, I would like to express my heartfelt appreciation for the opportunity to have my work published in this esteemed journal. The entire publication process was smooth and well-organized, and I am extremely satisfied with the final result. The Editorial Team demonstrated the utmost professionalism, providing prompt and insightful feedback throughout the review process. Their clear communication and constructive suggestions were invaluable in enhancing my manuscript, and their meticulous attention to detail and dedication to quality are truly commendable. Additionally, the support from the Editorial Office was exceptional. From the initial submission to the final publication, I was guided through every step of the process with great care and professionalism. The team's responsiveness and assistance made the entire experience both easy and stress-free. I am also deeply impressed by the quality and reputation of the journal. It is an honor to have my research featured in such a respected publication, and I am confident that it will make a meaningful contribution to the field.

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Dr Tewodros Kassahun Tarekegn

"I am grateful for the opportunity of contributing to [International Journal of Clinical Case Reports and Reviews] and for the rigorous review process that enhances the quality of research published in your esteemed journal. I sincerely appreciate the time and effort of your team who have dedicatedly helped me in improvising changes and modifying my manuscript. The insightful comments and constructive feedback provided have been invaluable in refining and strengthening my work".

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Dr Shweta Tiwari

I thank the ‘Journal of Clinical Research and Reports’ for accepting this article for publication. This is a rigorously peer reviewed journal which is on all major global scientific data bases. I note the review process was prompt, thorough and professionally critical. It gave us an insight into a number of important scientific/statistical issues. The review prompted us to review the relevant literature again and look at the limitations of the study. The peer reviewers were open, clear in the instructions and the editorial team was very prompt in their communication. This journal certainly publishes quality research articles. I would recommend the journal for any future publications.

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Dr Farooq Wandroo

Dear Jessica Magne, with gratitude for the joint work. Fast process of receiving and processing the submitted scientific materials in “Clinical Cardiology and Cardiovascular Interventions”. High level of competence of the editors with clear and correct recommendations and ideas for enriching the article.

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Dr Anyuta Ivanova

We found the peer review process quick and positive in its input. The support from the editorial officer has been very agile, always with the intention of improving the article and taking into account our subsequent corrections.

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Dr David Vinyes

My article, titled 'No Way Out of the Smartphone Epidemic Without Considering the Insights of Brain Research,' has been republished in the International Journal of Clinical Case Reports and Reviews. The review process was seamless and professional, with the editors being both friendly and supportive. I am deeply grateful for their efforts.

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Gertraud Teuchert-Noodt

To Dear Erin Aust – Editorial Coordinator of Journal of General Medicine and Clinical Practice! I declare that I am absolutely satisfied with your work carried out with great competence in following the manuscript during the various stages from its receipt, during the revision process to the final acceptance for publication. Thank Prof. Elvira Farina

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Dr Elvira Farina

Dear Jessica, and the super professional team of the ‘Clinical Cardiology and Cardiovascular Interventions’ I am sincerely grateful to the coordinated work of the journal team for the no problem with the submission of my manuscript: “Cardiometabolic Disorders in A Pregnant Woman with Severe Preeclampsia on the Background of Morbid Obesity (Case Report).” The review process by 5 experts was fast, and the comments were professional, which made it more specific and academic, and the process of publication and presentation of the article was excellent. I recommend that my colleagues publish articles in this journal, and I am interested in further scientific cooperation. Sincerely and best wishes, Dr. Oleg Golyanovskiy.

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Dr Oleg Golyanovski

Dear Ashley Rosa, Editorial Coordinator of the journal - Psychology and Mental Health Care. " The process of obtaining publication of my article in the Psychology and Mental Health Journal was positive in all areas. The peer review process resulted in a number of valuable comments, the editorial process was collaborative and timely, and the quality of this journal has been quickly noticed, resulting in alternative journals contacting me to publish with them." Warm regards, Susan Anne Smith, PhD. Australian Breastfeeding Association.

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Dr Susan Anne Smith

Dear Jessica Magne, Editorial Coordinator, Clinical Cardiology and Cardiovascular Interventions, Auctores Publishing LLC. I appreciate the journal (JCCI) editorial office support, the entire team leads were always ready to help, not only on technical front but also on thorough process. Also, I should thank dear reviewers’ attention to detail and creative approach to teach me and bring new insights by their comments. Surely, more discussions and introduction of other hemodynamic devices would provide better prevention and management of shock states. Your efforts and dedication in presenting educational materials in this journal are commendable. Best wishes from, Farahnaz Fallahian.

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Dr Farahnaz Fallahian

Dear Maria Emerson, Editorial Coordinator, International Journal of Clinical Case Reports and Reviews, Auctores Publishing LLC. I am delighted to have published our manuscript, "Acute Colonic Pseudo-Obstruction (ACPO): A rare but serious complication following caesarean section." I want to thank the editorial team, especially Maria Emerson, for their prompt review of the manuscript, quick responses to queries, and overall support. Yours sincerely Dr. Victor Olagundoye.

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Dr Victor Olagundoye

Dear Ashley Rosa, Editorial Coordinator, International Journal of Clinical Case Reports and Reviews. Many thanks for publishing this manuscript after I lost confidence the editors were most helpful, more than other journals Best wishes from, Susan Anne Smith, PhD. Australian Breastfeeding Association.

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Dr Susan Anne Smith

Dear Agrippa Hilda, Editorial Coordinator, Journal of Neuroscience and Neurological Surgery. The entire process including article submission, review, revision, and publication was extremely easy. The journal editor was prompt and helpful, and the reviewers contributed to the quality of the paper. Thank you so much! Eric Nussbaum, MD

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Dr Eric S Nussbaum

Dr Hala Al Shaikh This is to acknowledge that the peer review process for the article ’ A Novel Gnrh1 Gene Mutation in Four Omani Male Siblings, Presentation and Management ’ sent to the International Journal of Clinical Case Reports and Reviews was quick and smooth. The editorial office was prompt with easy communication.

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Hala Al Shaikh

Dear Erin Aust, Editorial Coordinator, Journal of General Medicine and Clinical Practice. We are pleased to share our experience with the “Journal of General Medicine and Clinical Practice”, following the successful publication of our article. The peer review process was thorough and constructive, helping to improve the clarity and quality of the manuscript. We are especially thankful to Ms. Erin Aust, the Editorial Coordinator, for her prompt communication and continuous support throughout the process. Her professionalism ensured a smooth and efficient publication experience. The journal upholds high editorial standards, and we highly recommend it to fellow researchers seeking a credible platform for their work. Best wishes By, Dr. Rakhi Mishra.

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Dr Rakhi Mishra

Dear Jessica Magne, Editorial Coordinator, Clinical Cardiology and Cardiovascular Interventions, Auctores Publishing LLC. The peer review process of the journal of Clinical Cardiology and Cardiovascular Interventions was excellent and fast, as was the support of the editorial office and the quality of the journal. Kind regards Walter F. Riesen Prof. Dr. Dr. h.c. Walter F. Riesen.

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Dr Walter F Riesen

Dear Ashley Rosa, Editorial Coordinator, International Journal of Clinical Case Reports and Reviews, Auctores Publishing LLC. Thank you for publishing our article, Exploring Clozapine's Efficacy in Managing Aggression: A Multiple Single-Case Study in Forensic Psychiatry in the international journal of clinical case reports and reviews. We found the peer review process very professional and efficient. The comments were constructive, and the whole process was efficient. On behalf of the co-authors, I would like to thank you for publishing this article. With regards, Dr. Jelle R. Lettinga.

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Dr Jelle Lettinga

Dear Clarissa Eric, Editorial Coordinator, Journal of Clinical Case Reports and Studies, I would like to express my deep admiration for the exceptional professionalism demonstrated by your journal. I am thoroughly impressed by the speed of the editorial process, the substantive and insightful reviews, and the meticulous preparation of the manuscript for publication. Additionally, I greatly appreciate the courteous and immediate responses from your editorial office to all my inquiries. Best Regards, Dariusz Ziora

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Dariusz Ziora

Dear Chrystine Mejia, Editorial Coordinator, Journal of Neurodegeneration and Neurorehabilitation, Auctores Publishing LLC, We would like to thank the editorial team for the smooth and high-quality communication leading up to the publication of our article in the Journal of Neurodegeneration and Neurorehabilitation. The reviewers have extensive knowledge in the field, and their relevant questions helped to add value to our publication. Kind regards, Dr. Ravi Shrivastava.

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Dr Ravi Shrivastava

Dear Clarissa Eric, Editorial Coordinator, Journal of Clinical Case Reports and Studies, Auctores Publishing LLC, USA Office: +1-(302)-520-2644. I would like to express my sincere appreciation for the efficient and professional handling of my case report by the ‘Journal of Clinical Case Reports and Studies’. The peer review process was not only fast but also highly constructive—the reviewers’ comments were clear, relevant, and greatly helped me improve the quality and clarity of my manuscript. I also received excellent support from the editorial office throughout the process. Communication was smooth and timely, and I felt well guided at every stage, from submission to publication. The overall quality and rigor of the journal are truly commendable. I am pleased to have published my work with Journal of Clinical Case Reports and Studies, and I look forward to future opportunities for collaboration. Sincerely, Aline Tollet, UCLouvain.

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Dr Aline Tollet

Dear Ms. Mayra Duenas, Editorial Coordinator, International Journal of Clinical Case Reports and Reviews. “The International Journal of Clinical Case Reports and Reviews represented the “ideal house” to share with the research community a first experience with the use of the Simeox device for speech rehabilitation. High scientific reputation and attractive website communication were first determinants for the selection of this Journal, and the following submission process exceeded expectations: fast but highly professional peer review, great support by the editorial office, elegant graphic layout. Exactly what a dynamic research team - also composed by allied professionals - needs!" From, Chiara Beccaluva, PT - Italy.

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Dr Chiara Giuseppina Beccaluva

Dear Maria Emerson, Editorial Coordinator, we have deeply appreciated the professionalism demonstrated by the International Journal of Clinical Case Reports and Reviews. The reviewers have extensive knowledge of our field and have been very efficient and fast in supporting the process. I am really looking forward to further collaboration. Thanks. Best regards, Dr. Claudio Ligresti

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Dr Claudio Ligresti

Dear Chrystine Mejia, Editorial Coordinator, Journal of Neurodegeneration and Neurorehabilitation. “The peer review process was efficient and constructive, and the editorial office provided excellent communication and support throughout. The journal ensures scientific rigor and high editorial standards, while also offering a smooth and timely publication process. We sincerely appreciate the work of the editorial team in facilitating the dissemination of innovative approaches such as the Bonori Method.” Best regards, Dr. Matteo Bonori.

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Dr Matteo Bonori

I recommend without hesitation submitting relevant papers on medical decision making to the International Journal of Clinical Case Reports and Reviews. I am very grateful to the editorial staff. Maria Emerson was a pleasure to communicate with. The time from submission to publication was an extremely short 3 weeks. The editorial staff submitted the paper to three reviewers. Two of the reviewers commented positively on the value of publishing the paper. The editorial staff quickly recognized the third reviewer’s comments as an unjust attempt to reject the paper. I revised the paper as recommended by the first two reviewers.

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Edouard Kujawski

Dear Maria Emerson, Editorial Coordinator, Journal of Clinical Research and Reports. Thank you for publishing our case report: "Clinical Case of Effective Fetal Stem Cells Treatment in a Patient with Autism Spectrum Disorder" within the "Journal of Clinical Research and Reports" being submitted by the team of EmCell doctors from Kyiv, Ukraine. We much appreciate a professional and transparent peer-review process from Auctores. All research Doctors are so grateful to your Editorial Office and Auctores Publishing support! I amiably wish our article publication maintained a top quality of your International Scientific Journal. My best wishes for a prosperity of the Journal of Clinical Research and Reports. Hope our scientific relationship and cooperation will remain long lasting. Thank you very much indeed. Kind regards, Dr. Andriy Sinelnyk Cell Therapy Center EmCell

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Dr Andriy Sinelnyk

Dear Editorial Team, Clinical Cardiology and Cardiovascular Interventions. It was truly a rewarding experience to work with the journal “Clinical Cardiology and Cardiovascular Interventions”. The peer review process was insightful and encouraging, helping us refine our work to a higher standard. The editorial office offered exceptional support with prompt and thoughtful communication. I highly value the journal’s role in promoting scientific advancement and am honored to be part of it. Best regards, Meng-Jou Lee, MD, Department of Anesthesiology, National Taiwan University Hospital.

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Dr Meng-JouLe

Dear Editorial Team, Journal-Clinical Cardiology and Cardiovascular Interventions, “Publishing my article with Clinical Cardiology and Cardiovascular Interventions has been a highly positive experience. The peer-review process was rigorous yet supportive, offering valuable feedback that strengthened my work. The editorial team demonstrated exceptional professionalism, prompt communication, and a genuine commitment to maintaining the highest scientific standards. I am very pleased with the publication quality and proud to be associated with such a reputable journal.” Warm regards, Dr. Mahmoud Kamal Moustafa Ahmed

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Mahmoud Kamal Moustafa Ahmed

Dear Maria Emerson, Editorial Coordinator of ‘International Journal of Clinical Case Reports and Reviews’, I appreciate the opportunity to publish my article with your journal. The editorial office provided clear communication during the submission and review process, and I found the overall experience professional and constructive. Best regards, Elena Salvatore.

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Dr Elena Salvatore

Dear Mayra Duenas, Editorial Coordinator of ‘International Journal of Clinical Case Reports and Reviews Herewith I confirm an optimal peer review process and a great support of the editorial office of the present journal

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Christoph Maurer

Dear Editorial Team, Clinical Cardiology and Cardiovascular Interventions. I am really grateful for the peers review; their feedback gave me the opportunity to reflect on the message and impact of my work and to ameliorate the article. The editors did a great job in addition by encouraging me to continue with the process of publishing.

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Baciulescu Laura

Dear Cecilia Lilly, Editorial Coordinator, Endocrinology and Disorders, Thank you so much for your quick response regarding reviewing and all process till publishing our manuscript entitled: Prevalence of Pre-Diabetes and its Associated Risk Factors Among Nile College Students, Sudan. Best regards, Dr Mamoun Magzoub.

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Dr Mamoun Magzoub

International Journal of Clinical Case Reports and Reviews is a high quality journal that has a clear and concise submission process. The peer review process was comprehensive and constructive. Support from the editorial office was excellent, since the administrative staff were responsive. The journal provides a fast and timely publication timeline.

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Joel Yat Seng Wong

Dear Maria Emerson, Editorial Coordinator of International Journal of Clinical Case Reports and Reviews, What distinguishes International Journal of Clinical Case Report and Review is not only the scientific rigor of its publications, but the intellectual climate in which research is evaluated. The submission process is refreshingly free of unnecessary formal barriers and bureaucratic rituals that often complicate academic publishing without adding real value. The peer-review system is demanding yet constructive, guided by genuine scientific dialogue rather than hierarchical or authoritarian attitudes. Reviewers act as collaborators in improving the manuscript, not as gatekeepers imposing arbitrary standards. This journal offers a rare balance: high methodological standards combined with a respectful, transparent, and supportive editorial approach. In an era where publishing can feel more burdensome than research itself, this platform restores the original purpose of peer review — to refine ideas, not to obstruct them Prof. Perlat Kapisyzi, FCCP PULMONOLOGIST AND THORACIC IMAGING.

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Dr Perlat Kapisyzi

Dear Grace Pierce, International Journal of Clinical Case Reports and Reviews I appreciate the opportunity to review for Auctore Journal, as the overall editorial process was smooth, transparent and professionally managed. This journal maintains high scientific standards and ensures timely communications with authors, which is truly commendable. I would like to express my special thanks to editor Grace Pierce for his constant guidance, promt responses, and supportive coordination throughout the review process. I am also greatful to Eleanor Bailey from the finance department for her clear communication and efficient handling of all administrative matters. Overall, my experience with Auctore Journal has been highly positive and rewarding. Best regards, Sabita sinha

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Sabita sinha

Dear Mayra Duenas, Editorial Coordinator of the journal IJCCR, I write here a little on my experience as an author submitting to the International Journal of Clinical Case Reports and Reviews (IJCCR). This was my first submission to IJCCR and my manuscript was inherently an outsider’s effort. It attempted to broadly identify and then make some sense of life’s under-appreciated mysteries. I initially had responded to a request for possible submissions. I then contacted IJCCR with a tentative topic for a manuscript. They quickly got back with an approval for the submission, but with a particular requirement that it be medically relevant. I then put together a manuscript and submitted it. After the usual back-and-forth over forms and formality, the manuscript was sent off for reviews. Within 2 weeks I got back 4 reviews which were both helpful and also surprising. Surprising in that the topic was somewhat foreign to medical literature. My subsequent updates in response to the reviewer comments went smoothly and in short order I had a series of proofs to evaluate. All in all, the whole publication process seemed outstanding. It was both helpful in terms of the paper’s content and also in terms of its efficient and friendly communications. Thank you all very much. Sincerely, Ted Christopher, Rochester, NY.

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Dr Ted Christopher