Evaluation of levetiracetam for early post-traumatic seizure prophylaxis: A level II trauma center experience.

INTRODUCTION: Traumatic brain injury (TBI) can induce early or late post-traumatic seizures (PTS). While PTS incidence is low, prophylaxis is used despite a lack of consensus on agent or duration. Levetiracetam (LEV) for early PTS prophylaxis is preferred due to its safety and efficacy. The purpose of this study was to evaluate LEV for early PTS prophylaxis. METHODS AND MATERIALS: A single-center, retrospective chart review of TBI patients ≥18 years who received LEV for early PTS prophylaxis between August 2018-July 2019. The primary outcome was LEV duration. Secondary outcomes were incidence of seizure, intensive care unit (ICU) and hospital length of stay (LOS). RESULTS: Of the 137 included, mean age was 59 ± 20 years and 69.3% were male. The mean admission GCS was 13 ± 4 and 77.4% had mild TBI. Median LEV duration was 7 (IQR 4-10) days and 13.9% met recommended 7-day duration. Those prescribed LEV >7 days had more than twice the median LEV duration than those prescribed ≤7 days [10.25 (8.5-15.5) vs 4 (1.5-4.5) days, p < 0.0001]. Electroencephalography-confirmed PTS occurred in 2.2%, with an early PTS incidence of 0.73%. Median ICU and hospital LOS were 2 (IQR 1-7) and 7 (IQR 3-16) days, respectively. CONCLUSIONS: The incidence of PTS was low as most patients in our study had mild or moderate TBI. Early PTS prophylaxis with LEV for 7 days is appropriate, although the majority of patients did not meet the recommended duration. Efforts to standardize and implement PTS prophylaxis protocols are needed.

MitoFlex: an efficient, high-performance toolkit for animal mitogenome assembly, annotation and visualization.

SUMMARY: MitoFlex is a linux-based mitochondrial genome analysis toolkit, which provides a complete workflow of raw data filtering, de novo assembly, mitochondrial genome identification and annotation for animal High Throughput Sequencing data. The overall performance was compared between MitoFlex and its analogue MitoZ, in terms of protein-coding gene recovery, memory consumption and processing speed. AVAILABILITYAND IMPLEMENTATION: MitoFlex is available at https://github.com/Prunoideae/MitoFlex under GPLv3 license. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

A New Species and New Record of Freshwater Turbellarians of the Genus Gieysztoria (Platyhelminthes: Rhabdocoela) from China.

A new species of Dalyelliidae, Gieysztoria pellucida Wang and You, is described based on material collected in southern China through an integrative approach combining morphological, histological, and molecular (18S and 28S rDNA) data. Gieysztoria pellucida sp. nov. is morphologically characterized by a fan-shaped (about 270° when pressed) stylet, consisting of 13 similar distal spines and a broad girdle without fenestrae region. This stylet is distinct from that of any other similar species in the Aequales group to which this species belongs. In addition, specimens identifiable as Gieysztoria garudae Van Steenkiste, Van Mulken, and Artois, 2012 were discovered from the same location as G. pellucida sp. nov. Gieysztoria garudae has previously been known only from India; the present study thus represents the first record of the species from China.

Two New Brackish-Water Species of Macrostomum (Platyhelminthes: Macrostomorpha) from China and Their Phylogenetic Positions.

In this paper, two new brackish-water species of the macrostomid turbellarian genus Macrostomum, Macrostomum pseudosinense sp. nov. and Macrostomum taurinum sp. nov., collected from coastal water at Shenzhen, Guangdong Province, China, are described based on morphological, histological, and molecular phylogenetic analyses. Macrostomum pseudosinense sp. nov. differs from similar species within the genus in the length of the stylet (152 ± 15.0 µm), diameter of stylet opening (20 ± 4.0 µm proximally; 7 ± 0.5 µm distally), two bends of the stylet, and the non-spiral end of the stylet. Macrostomum taurinum sp. nov. differs from its congeners in the length of the stylet (81 ± 7.4 µm), the stylet bending position and angle (50% and 60°), diameter of stylet proximal opening (15 ± 3.0 µm), sperm with bristles and brush, and the smooth-walled ovaries. Phylogenetic analyses inferred from nuclear 18S and 28S rRNA genes support the establishments of these two new species. In addition, reciprocal mating behavior of M. pseudosinense sp. nov. was observed and documented.

Mechanisms of thermal treatment on two dominant copepod species in O3/BAC processing of drinking water.

Phyllodiaptomus tunguidus and Heliodiaptomus falxus are dominant copepods species in drinking water processing plants in southern China. With a potential penetration risk, the breeding and leakage of copepods are drawing more and more attention in recent years. The current study provided a thermal treatment method to control copepods and their eggs. Results showed that: (1) the immediate death rates of P. tunguidus and H. falxus after heated to 34-40 °C for 5 min are positively correlated to the treatment temperatures (P < 0.01), and all individuals of the both species were eliminated after heated at 40 °C for 5 min; (2) overall hatching rates of P. tunguidus eggs were negatively correlated with treatment temperatures (P < 0.01) between 39-45 °C, with zero percent hatched after treatment at 45 °C for 5 min; (3) hatching rates of H. falxus were negatively correlated with treatment temperatures (P < 0.01) between 37-41 °C, with no nauplii hatched when treated at 41 °C for 5 min; (4) paraffin section histological examination indicated that thermal treatment caused severe damage to internal organs and egg structure. Finally, based on the experimental data, the application of the thermal treatment method was discussed in ozonation combined with biological activated carbon (O3/BAC) processing of drink water treatment.

Increased expression of tumor proliferation genes in Hispanic women with early-stage breast cancer.

Hispanic women have higher breast cancer mortality compared to non-Hispanic whites. We evaluated for Proliferation Axis Score differences, as determined by Oncotype Dx, in Hispanic and non-Hispanic white women with newly diagnosed breast cancer. We matched 219 women, based upon age, stage, and nodal status. Compared to non-Hispanic whites, Hispanic women with hormone-sensitive, HER2-negative early-stage breast cancer had a higher Proliferation Axis Score. No differences were seen in Recurrence Score, ER, PR, or HER2 by Oncotype DX. CCNB1 and AURKA were significantly higher in Hispanic women. These tumor differences may help explain breast cancer outcome differences between the two ethnicities.

Presurgical trial of metformin in overweight and obese patients with newly diagnosed breast cancer.

INTRODUCTION: We conducted a presurgical trial to assess the tissue-related effects of metformin in overweight/obese breast cancer (BC) patients. METHODS: Metformin 1,500 mg daily was administered to 35 nondiabetics with stage 0-III BC, body mass index (BMI) ≥ 25 kg/m(2). The primary endpoint was tumor proliferation change (i.e., ki-67). Tumor proliferation change was compared to untreated historical controls, matched by age, BMI, and stage. RESULTS: There was no reduction in ln(ki-67) after metformin (p = .98) or compared to controls (p = .47). There was a significant reduction in BMI, cholesterol, and leptin. CONCLUSION: Despite no proliferation changes, we observed reductions in other relevant biomarkers.

Integrative description of a new species of Dugesia (Platyhelminthes, Tricladida, Dugesiidae) from Shennongjia, Central China.

A new species of the genus Dugesia (Platyhelminthes, Tricladida, Dugesiidae) from Xiangxi River, Shennongjia Forestry District, Hubei Province, China, is described on the basis of an integrative approach, involving morphology, and molecular systematics. The new species Dugesia saccaria A-T. Wang & Sluys, sp. nov. is characterized by the following features: a dumb-bell-shaped, muscularized hump located just anterior to the knee-shaped bend in the bursal canal; a ventrally displaced ejaculatory duct, which, however, opens terminally through the dorsal portion of the blunt tip of the penis papilla; a ventrally located seminal vesicle, giving rise to a vertically running duct that eventually curves downwards to communicate with the ejaculatory duct via a small diaphragm; oviducts opening asymmetrically into the dorsal portion of the common atrium and at the knee-shaped part of the bursal canal. The phylogenetic position of the new species was determined using four molecular markers (18S rDNA; ITS-1; 28S rDNA; COI), which suggested that it groups with other species of Dugesia from the Australasian and Oriental biogeographical regions.

Inhibition of the PI3K/AKT pathway potentiates cytotoxicity of EGFR kinase inhibitors in triple-negative breast cancer cells.

Triple-negative breast cancers (TNBCs) are known to be intrinsically resistant to inhibitors for epidermal growth factor receptor (EGFR). Until now, clinical trials for TNBCs using EGFR inhibitors (EGFRis) as single agents have yielded disappointing results. Here, we report that combinatorial treatment using EGFRis, such as gefitinib or erlotinib, with PI3K/AKT pathway inhibitors (PI3K/AKTis) demonstrated a synergistic, anti-proliferative effect in cell lines of the basal-like (BL) subtype, a subtype of TNBC. Western blot analysis revealed that the gefitinib/PI-103 combination significantly reduced the level of both phospho-AKT and phospho-ERK in two susceptible BL subtype cell lines, SUM149PT and MDA-MB-468, whereas it had little or no effect on the level of phospho-ERK in two non-susceptible cell lines (HS578T and MDA-MB-231) of mesenchymal stem-like (MSL) TNBC subtype. The gefitinib/PI-103 combination also significantly induced caspase-3/7-mediated PARP cleavage and reduced two anti-apoptotic proteins, XIAP and Bcl-2 in the susceptible cell lines. In addition, the level of myeloid cell leukemia 1 (Mcl-1) protein was markedly decreased by gefitinib/PI-103 combination in the BL TNBC cells, but showed no significant change by this combination in MSL subtype cells. These results suggest that pharmacological inhibition of EGFR used in combination of PI3K/AKTis is a potential therapeutic approach to treat a subtype of TNBCs.

Inhibition of checkpoint kinase 2 (CHK2) enhances sensitivity of pancreatic adenocarcinoma cells to gemcitabine.

Checkpoint kinase 2 (CHK2) plays pivotal function as an effector of cell cycle checkpoint arrest following DNA damage. Recently, we found that co-treatment of NSC109555 (a potent and selective CHK2 inhibitor) potentiated the cytotoxic effect of gemcitabine (GEM) in pancreatic cancer MIA PaCa-2 cells. Here, we further examined whether NSC109555 could enhance the antitumour effect of GEM in pancreatic adenocarcinoma cell lines. In this study, the combination treatment of NSC109555 plus GEM demonstrated strong synergistic antitumour effect in four pancreatic cancer cells (MIA PaCa-2, CFPAC-1, Panc-1 and BxPC-3). In addition, the GEM/NSC109555 combination significantly increased the level of intracellular reactive oxygen species (ROS), accompanied by induction of apoptotic cell death. Inhibition of ROS generation by N-acetyl cysteine (NAC) significantly reversed the effect of GEM/NSC109555 in apoptosis and cytotoxicity. Furthermore, genetic knockdown of CHK2 by siRNA enhanced GEM-induced apoptotic cell death. These findings suggest that inhibition of CHK2 would be a beneficial therapeutic approach for pancreatic cancer therapy in clinical treatment.

Combined histone deacetylase and cyclooxygenase inhibition achieves enhanced antiangiogenic effects in lung cancer cells.

Prostaglandin E2 (PGE2) is an important pro-angiogenic and pro-proliferative cytokine and the key enzymes modulating its levels, cyclooxygenase (COX)-2 and 15-hydroxyprostaglandin dehydrogenase (15-PGDH) play important opposing roles in carcinogenesis. Previously we found loss of 15-PGDH expression in lung cancer and its reactivation leads to strong in vivo tumor-suppressive effect via an antiangiogenic mechanism. Here, we find that HDAC inhibitors (HDACI), such as trichostatin A (TSA) and vorinostat could reactivate 15-PGDH expression but overall induce PGE2 generation and this is the result of concomitant induction of COX-1 and -2 leading to functional promotion of endothelial cell proliferation and capillary formation. Direct TSA treatment inhibits endothelial cell proliferation and capillary formation in our study in line with prior reports as HDACIs have been shown to directly inhibit angiogenesis. The elevation of PGE2 levels induced by HDACI is potently neutralized by indomethacin (INN) or Celecoxib co-treatment and accordingly, angiogenesis is more effectively inhibited when using conditioned medium of co-treatment than either alone confirming that this effect is mediated via the PGE2 axis. Accordingly, blockage of EP2/4 receptors mitigates the stimulation of angiogenesis by excessive PGE2 generation mediated by TSA. In this study, we identify a potentially adverse effect of HDACIs through induction of both 15-PGDH and COX-2 leading to elevated PGE2 levels and thereby stimulation of angiogenesis. Co-treatment of TSA and INN shows more potent anti-angiogenic effects by inducing 15-PGDH and inhibiting COX-2. Overall, our results suggest that combined HDACI and COX inhibition should be explored clinically to achieve more meaningful benefits from HDACI therapy in lung cancer.

Inhibition of constitutively activated phosphoinositide 3-kinase/AKT pathway enhances antitumor activity of chemotherapeutic agents in breast cancer susceptibility gene 1-defective breast cancer cells.

Loss or decrease of wild type BRCA1 function, by either mutation or reduced expression, has a role in hereditary and sporadic human breast and ovarian cancers. We report here that the PI3K/AKT pathway is constitutively active in BRCA1-defective human breast cancer cells. Levels of phospho-AKT are sustained even after serum starvation in breast cancer cells carrying deleterious BRCA1 mutations. Knockdown of BRCA1 in MCF7 cells increases the amount of phospho-AKT and sensitizes cells to small molecule protein kinase inhibitors (PKIs) targeting the PI3K/AKT pathway. Restoration of wild type BRCA1 inhibits the activated PI3K/AKT pathway and de-sensitizes cells to PKIs targeting this pathway in BRCA1 mutant breast cancer cells, regardless of PTEN mutations. In addition, clinical PI3K/mTOR inhibitors, PI-103, and BEZ235, showed anti-proliferative effects on BRCA1 mutant breast cancer cell lines and synergism in combination with chemotherapeutic drugs, cisplatin, doxorubicin, topotecan, and gemcitabine. BEZ235 synergizes with the anti-proliferative effects of gemcitabine by enhancing caspase-3/7 activity. Our results suggest that the PI3K/AKT pathway can be an important signaling pathway for the survival of BRCA1-defective breast cancer cells and pharmacological inhibition of this pathway is a plausible treatment for a subset of breast cancers.

MicroRNA deregulation in polycythemia vera and essential thrombocythemia patients.

Polycythemia vera (PV) and essential thrombocythemia (ET) are the two most common myeloproliferative neoplasms. The same JAK2(V617F) mutation can be found in both disorders and is able to recapitulate many of the phenotypic abnormalities of these diseases in the murine models. The disease phenotype is also influenced by other unknown genetic or epigenetic factors. MicroRNAs (miRNA) are 18-24 nucleotides single-stranded non-protein-coding RNAs that function primarily as gene repressors by binding to their target messenger RNAs. We performed miRNA expression profiling by oligonucleotide microarray analysis in purified peripheral blood CD34+ cells from eight JAK2(V617F)-positive PV patients and six healthy donors. A quantitative reverse-transcription polymerase chain reaction assay was used to verify differential miRNA expression. Since erythrocytosis is the only feature that distinguishes PV from ET, we also compared specific miRNA expression in the nucleated erythroid cells directly descended from the early erythroid progenitor cells of PV and ET patients. Our data indicate that significant miRNA deregulation occurs in PV CD34+ cells and confirm a genetic basis for the gender-specific differences that characterize PV with respect to miRNA. The results of our study also suggest that deregulated miRNAs may represent an important mechanism by which the PV erythrocytosis and ET thrombocytosis phenotypes are determined.

Thymidylate synthase expression and molecular alterations in adenosquamous carcinoma of the lung.

Thymidylate synthase expression is known to be higher in squamous cell carcinoma than in adenocarcinoma of the lung. It is thought that this is the reason for the poor efficacy of pemetrexed in squamous cell carcinoma. However, there is limited data on thymidylate synthase expression in adenosquamous carcinoma, a distinct subtype of lung cancer containing both squamous and glandular differentiation. Furthermore, molecular alterations like epidermal growth factor receptor and Kirsten rat sarcoma 2 viral oncogene homolog mutations, which are seen in adenocarcinomas, are not well understood in mixed histology tumors such as adenosquamous carcinoma. In our study, we sought to better characterize adenosquamous tumors of the lung. Using immunohistochemistry to evaluate thymidylate synthase protein levels, we found that the expression of thymidylate synthase in these mixed tumors roughly parallel that of squamous cell carcinoma, instead of falling in between squamous cell and adenocarcinoma. Of note, in adenosquamous samples, the expression of thymidylate synthase was more closely correlated within the two components than would be expected by random chance alone. Also, we had a relatively high rate of epidermal growth factor receptor (11%) and Kirsten rat sarcoma 2 viral oncogene homolog (33%) mutations in these specimens, with the mutations showing convergence in both the glandular and squamous components upon microdissection. Our results indicate that adenosquamous carcinomas are not simple mixtures of their two histological components; they rather behave as their own entity, and it is important to further understand their behavior. Given the similarity of thymidylate synthase expression between squamous cell and adenosquamous carcinoma, and that thymidylate synthase is the main target of pemetrexed, we extrapolate that pemetrexed may also have inferior clinical activity in adenosquamous carcinoma.

An evolving paradigm for the workup and management of high-risk cutaneous squamous cell carcinoma.

BACKGROUND: No established standard of care exists for aggressive cutaneous squamous cell carcinoma (CSCC). OBJECTIVE: We sought to establish an aggressive CSCC management protocol by reviewing high-risk CSCC (HCSCC) and very high-risk CSCC (VCSCC) cases at our institution. METHODS: This was a retrospective review of all CSCC cases treated at our institution. RESULTS: A total of 27 patients were identified of 1591 cases treated between 2000 and 2011. Four patients with HCSCC received surgery alone and 1 received surgery and radiation. All remain disease free (median follow-up 5 years). Among patients with VCSCC, 4 received surgery alone: 1 (25%) showing a complete response and 3 (75%) showing disease progression. Eleven received surgery and radiation: 4 (36.4%) with complete response (median follow-up 3 years) and 7 (63.6%) with disease progression (median time to recurrence 6 months). Six received surgery and cetuximab: 3 (50%) had a complete response (median follow-up 3 years), 2 (33%) had disease progression, and 1 (14%) could not be assessed because of inability to tolerate infusions. One patient received surgery, cetuximab, and radiation, and remains disease-free after 4 years. LIMITATIONS: Lack of randomization, blinding, a true control arm, or standardization of treatment protocols are limitations. CONCLUSIONS: Patients with very HCSCC may have improved outcomes with surgery and adjuvant cetuximab.

Two new species of Gieysztoria (Platyhelminthes, Rhabdocoela, Dalyelliidae) from a freshwater artificial lake in Shenzhen, China.

Two new species of the "Aequales" of genus Gieysztoria were collected and described from an artificial lake on the Shenzhen University campus. Gieysztoria bimaculata n. sp., is distinguished based on two groups clavate pigmentations dorsally between the pharynx and intestine, and has a sclerotic stylet comprising a proximal girdle with 40-46 distal dagger-shape spines, thus has the maximum number of spines within "Aequales" group. Gieysztoria guangdongensis n. sp. has a sclerotic stylet with a proximal girdle and 18 distal blade-shaped spines. Comparison with similar species based mainly on stylet morphology suggests that Gieysztoria bimaculata n. sp. and Gieysztoria guangdongensis n. sp. are apparently different from the known species of Gieysztoria in this moment. In addition, the stability of the amount of distal spines of "Aequales" species is briefly discussed.

Effect of Alternatives to Opiates Program on Discharge Opioid Prescribing in Trauma Patients.

BACKGROUND: Opioid overdose deaths have increased over the last two decades, despite efforts to reduce prescribing. This study aimed to determine if a hospital-wide Alternatives to Opiates (ALTOSM) program reduced opioid prescribing in hospital and upon discharge after trauma. OBJECTIVES: The primary outcome was incidence of opioid prescribing at hospital discharge Pre- and Post-ALTO. Secondary outcomes were the percent of patients with in-hospital opioid, non-opioid and multimodal analgesia, and hospital and intensive care unit (ICU) length of stay (LOS). METHODS: This is a single-center, retrospective analysis of patients >/ = 18 years old admitted for >24 hours with the primary diagnosis of traumatic injury between August 2018 - October 2019. Patients with alcohol or polysubstance abuse, chronic opioid use, or in-hospital mortality were excluded. RESULTS: A total of 703 patients were included, 471 in Pre-ALTO and 232 in Post-ALTO groups. The mean age was 59 ± 22 years and most were male (58.7%). Mean initial Injury Severity Score (ISS) was 9.1 ± 7.7. Opioid prescribing at hospital discharge occurred more in the Post-ALTO group (132/332, 39.4% vs 90/203, 43.8%; P = .1237). Most patients were prescribed in-hospital opioid (332/471, 70.4% vs 203/232, 87.5%, P < .0001) and non-opioid (441/471, 93.6% vs 229/232, 98.7%; P = .0027) analgesics, or multimodal analgesia (397/471, 84.3% vs 203/232, 87.5%; P = .2591). Median hospital and ICU LOS were also similar between groups [5 (3-9) vs 4(3-7), P = .3427] and ICU [2(0-4) vs 3(2-5), P = .3461]. CONCLUSION: Opioids remain mainstay for trauma-related pain treatment. ALTOSM was not associated with less in-hospital or discharge opioid prescribing.

Opioid Prescribing at Discharge in Opioid-Naïve Trauma Patients.

BACKGROUND: Opioid analgesics remain mainstay of treatment for trauma-related pain despite growing concerns for opioid dependency or misuse. The purpose of this study was to evaluate opioid prescribing at hospital discharge after traumatic injury. METHODS: This is a single-center, retrospective analysis of patients ≥18 years of age admitted for ≥24 hours with a primary diagnosis of traumatic injury. Those with alcohol use disorder, polysubstance abuse, chronic opioid use, or in-hospital mortality were excluded. The primary outcome was the incidence of patients prescribed opioids at discharge. Secondary outcomes included percent of patients who received nonopioids, intensive care unit (ICU) admission, and hospital length of stay (LOS). RESULTS: Of the 927 encounters, 471 were included. The mean age was 60 ± 23 years, and 62.0% were male. The majority were blunt trauma, and 49.9% were falls. Mean initial injury severity score (ISS) was 9 ± 7.2. Of the 70.4% of patients prescribed opioids, 39.4% were discharged on opioids. Age ≥30 years, ICU admission, ISS <9, or Charlson Comorbidity Index >1 was less likely to have opioids prescribed at discharge. Most received nonopioids (93.6%) and multimodal analgesia (84.3%). The median hospital and ICU LOS were 5 (3-9) and 2 (0-4) days, respectively. DISCUSSION: Only 39.4% had opioids prescribed at discharge. Opioid-reductive strategies may decrease in-hospital and discharge opioid prescribing. While opioid analgesics remain a mainstay of trauma-associated pain management, institution-wide opioid-sparing strategies can further reduce discharge opioid prescribing after trauma.

A new species of hydra (Cnidaria: Hydrozoa: Hydridae) and molecular phylogenetic analysis of six congeners from China.

A new species of genus Hydra (Cnidaria: Hydrozoa: Hydridae), Hydra shenzhensis sp. nov. from Guangdong Province, China, is described and illustrated. Most polyps have five tentacles. Column length reaches 11 mm when relaxed. Buds do not acquire tentacles synchronously. Stenotele is broad and pyriform in shape, 1.2 times as long as its width. Holotrichous isorhiza is asymmetrical and slender (more than 2.7 times as long as its width), with transverse and slanting coils. Atrichous isorhiza is long, resembling a melon-seed in shape. Desmoneme is asymmetrically pyriform in shape. The new species, belonging to the vulgaris group, is dioecious; sexual reproduction was found to occur mostly during November and December under conditions of dense culture or food shortage. Two to thirteen testes, cone-like shape with papilla, formed beneath the tentacles. One to three ovaries, with an egg cup, milky white in color, formed on body column. Ninety percent of individuals developed only one ovum. On a mother polyp, a fertilized ovum developed an embryonic theca covering its surface. The embryotheca is brown, with a spine-like structure, covering a layer of transparent, membrane-like material. For phylogenetic analysis, the mitochondrial cytochrome oxidase subunit I gene (COI) of six hydra species collected from China was amplified by polymerase chain reaction (PCR) and sequenced. Morphological characters in combination with molecular evidence support the hydra described here as a new species.

Three new species of Macrostomum (Platyhelminthes, Macrostomorpha) from China and Australia, with notes on taxonomy and phylogenetics.

In this paper, three species of the macrostomid free-living flatworm genus Macrostomum are described. Two species, Macrostomumlittorale Wang & Shi, sp. nov. and M.shekouense Wang & Shi, sp. nov., were collected from coastal water at Shenzhen, Guangdong Province, China. One species, M.brandi Wang & Shi, sp. nov., was collected from Perth, Western Australia and Queenscliff, Victoria, Australia. Macrostomumlittorale sp. nov. differs from congeneric species within the genus in the length of the stylet, diameter of stylet opening, and the bend of the stylet. Macrostomumshekouense sp. nov. and M.brandi sp. nov. differ from similar species within the genus in the stylet morphology, position of the female antrum and developing eggs, or presence or absence of the false seminal vesicle. Phylogenetic analysis based on cytochrome c oxidase subunit I (COI) gene shows that M.littorale sp. nov. and M.hystrix are sister clades on two well-separated branch, M.shekouense sp. nov. and M.brandi sp. nov. are sister clades on two well-separated branches. Accordingly, both morphological and molecular evidence support M.littorale sp. nov., M.shekouense sp. nov., and M.brandi sp. nov. as three new species.