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We aim to describe the basic demographics, clinical course and outcomes of critically ill patients with Covid-19 admitted to Avera McKennan Hospital and University Health Center Intensive Care Unit (ICU) between March 20 and May 4, 2020.In this single centered, retrospective, observational study, we enrolled 37 critically ill adults with COVID-19 pneumonia admitted to the (ICU) between March 20 and May 4, 2020. Demographic data, admitting symptoms, laboratory values, co-morbidities, treatments and clinical outcomes were collected. Data was compared between survivors and non-survivors. We aim to describe our data and report the 28-day mortality as of June 1, 2020.Of 154 patients admitted with COVID-19 pneumonia during our study period, 37 (24 percent) were critically ill and required an ICU stay. The mean age was 58 years and 76 percent were men. Of these 37 patients, 28 (78 percent) had a chronic illness (diabetes in 43 percent, hypertension in 47 percent). In addition, 54 percent were associated with a local meat packing plant. Most common presenting symptoms were dyspnea (92 percent), cough (70 percent) and fever (68 percent). The mean PaO2/ FiO2 ratio was 143 (67-362). Significant lab findings include the following: 54 percent of patients had lymphocytopenia, the mean ferritin was 850 ng/mL (10-3528), the mean D-Dimer was 4.09 FEU ug/mL and the mean IL-6 was 96.5 pg/mL. At 28 days, 24 percent (nine) had died. Twenty-five (68 percent) patients required mechanical ventilation, with 10 (27 percent) of those patients requiring initiation of neuromuscular blocking agents for ventilator compliance. Of those four (40 percent) did not survive. In addition, 20 patients (54 percent) were proned. Pneumomediastinum or pneumothorax occurred in five of the 37 (14 percent). Renal replacement therapy was required in 6 of the 37 patients, 4 of whom (66 percent) died. Steroids were used in 70 percent of patients, tocilizumab in 59 percent, and hydroxychloroquine in 27 percent. All patients received antibiotics. Convalescent plasma became available for our 5th patient. A total of 29 (78 percent) received convalescent plasma, (86 percent of survivors and 56 percent non-survivors). Median ICU length of stay was 11 days for both survivors (1-49) and non-survivors (1-21). There were no differences in age, body mass index (BMI), or initial PaO2/FiO2 (P/F) among those two groups. Non-survivors (nine) included the two immune compromised patients in our cohort, two patients with pre-existing DNR/DNI status, and one death within two hours of admit. Compared with survivors, more of the non-survivors received vasopressors (78 percent vs 46 percent), dialysis (44 percent vs 7 percent) and hydroxychloroquine (44 percent vs 21 percent). The first 5 patients treated in the ICU did not survive. One month after the initial case was reported in South Dakota, our ICU experienced a six-week surge. At its highest, COVID-19-related census reached 63 percent of the ICU capacity (15/24).Mortality of critically ill patients with COVID-19 is high. Multi-organ, advanced and prolonged critical care resources are needed. Interpretation of our data is limited by a higher mortality of the earlier members of the cohort, a change in therapeutic practice over time and institution of social distancing.

Bluetongue (BT), caused by bluetongue virus (BTV), is a vector-borne disease of small ruminants that has the potential to spread across international borders. Despite large populations of susceptible animals and borders with BTV endemic countries, little is known of the disease burden and prevalent serotypes in the province of Balochistan in Pakistan. We conducted a cross-sectional study to determine seroconversion and prevalent serotypes in selected districts of the province using a competitive enzyme-linked immunosorbent assay (cELISA) and real-time polymerase chain reaction (RT-PCR). Sera (n = 876) were collected from clinically healthy sheep and goats originating from the districts of Quetta (n = 300), Mastung (n = 201), Killa Saifullah (n = 75) and Kech (n = 300). None of the study herds (n = 97) were seronegative for BTV, and at the individual level, the overall prevalence of BTV seroconversion was 47.26% (n = 414/876, 95% CI = 43.92%-50.63%). A higher percentage of goats (50.87%, 95% CI = 45.99%-55.73%) were seropositive for anti-VP7 immunoglobulins (IgG) than sheep (44.21%, 95% CI = 39.81%-48.70%). Odds ratios of seroconversion for goats were associated with breed type (χ2 = 16.84, p = .01), parity (χ2 = 23.66, p = .00) and presence of vector (χ2 = 2.63, p = .10), whereas for sheep, it was associated with breed type (χ2 = 13.80, p = .01) and parity (χ2 = 53.40, p = .00). Serotype 8 was the most prevalent (26.82%, 95% CI = 14.75%-43.21%) followed by an equal prevalence of serotypes 2 and 9 (7.31%, 95% CI = 1.91%-21.01%). To the best of our knowledge, this is the first study conducted in Balochistan province and the results indicate that there is a necessity to initiate intervention strategies to control BT disease burden not only in this region of Pakistan but also in adjacent areas of the neighbouring countries, Iran and Afghanistan.

Newcastle disease virus (NDV) live vaccines are supplied in lyophilized form and usually administered through conventional routes (drinking water, spray, or eye drop) following reconstitution in a diluent. Virus inactivation due to physico-chemical properties of the diluent at the time of administration may lead to vaccine failure. The present study aimed to evaluate the survival of NDV live vaccine strain immersed in 5 pH-amended water samples (pH 5.00, pH 6.00, pH 7.00, pH 8.00, and pH 9.00) by sequential determination of virus infectivity on Vero cells for 3 hours. Minimum reduction in virus infectivity was recorded in the water with neutral or slightly alkaline pH, while the virus was relatively less stable at extreme pH conditions. Maximum reduction of infectivity was observed in the water with pH 9.00 in which the virus was completely inactivated within 3 hours. Addition of stabilizers (Cevamune® or skimmed milk) slightly altered the pH and total dissolved solids (TDS) values of the virus-charged water samples. In the stabilizer-added water samples, minimum reduction in infectivity was observed in the water with neutral pH, followed by the ones with a pH of 8.00, 6.00, 5.00, and 9.00. In all types of water samples, T-90 values (time required for 90% reduction in virus infectivity) were highest (485 minutes) at neutral pH (pH 7.00) and lowest (102 to 134 min) at an extreme alkaline condition (pH 9.00). Results of the present study indicate that water with a pH range of 7.00 to 8.00 is suitable for administration of NDV live vaccines. However, the addition of Cevamune® or skimmed milk may have beneficial effects on preserving the infectivity of the virus, even at extreme pH conditions.

Copper nanoparticles (Cu NPs) have been synthesized by using laser ablation method, using deionized water as main solvent. The formation of Cu NPs is confirmed by UV–visible spectrophotometer (UV–Vis), atomic force microscopy (AFM) and X-ray diffraction (XRD). Cu NPs fabricated by laser ablation have diameter in the range from 14 to 55 nm. Structural analysis revealed the face-centered cubic (fcc) crystal structure of Cu NPs. The antibacterial activity of Cu NPs has been evaluated in vitro against strains of Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus). The fabricated Cu NPs show considerable antibacterial activity against both bacterial strains. The bacterial activity of Cu NPs was found to depend on the microbial species.

A novel contaminant was isolated from Madin Darby Bovine Kidney (MDBK) cells. The organism was unable to grow on standard microbiological media by conventional techniques, but grew well in Dulbecco's Modified Eagle's Medium (DMEM) containing high glucose concentration. The organism formed a white biofilm on the bottom without any signs of turbidity. Upon genome sequence analysis of 16 S rDNA, the contaminant was identified as Sphingomonas sp. Shah, a member of the group α-Proteobacteria. Neutral red dye uptake method confirmed clear cytotoxic potential of the bacterium on A-549 cells. The organism was capable of invading and infecting different mammalian cell lines: MDBK, ZZ-R, 293-T, A549, and HeLa cells. Infected cells showed a variety of cytopathic effects including vacuolation at perinuclear area, cytoplasmic granulation and membrane blebbing. Microscopic analysis of the infected cells revealed the presence of cytoplasmic vacuoles harboring motile organisms. Apparently local serum preparations seem to be the source of this contamination, which is imperceptibly passed on from one culture passage to the other and ultimately leading to serious cytopathic manifestations. J. Cell. Biochem. 116: 934–942, 2015. © 2015 Wiley Periodicals, Inc.

Objective: To determine the role of water fleas in accumulating avian influenza viruses (AIV) from the surrounding water and to estimate their role as a vector of AIV. Methods: Water fleas were exposed to H4N6 and H5N1 AIV-contaminated water in a closed system. The potential of water fleas to take up and retain the viruses was estimated by quantitative real-time RT-PCR (qRRT-PCR) and titration on cell culture. Results: Contamination trials showed that significantly higher amounts of viral RNA were detectable per gram of water fleas as compared to per milliliter of the surrounding water at 1, 4, and 6 days of incubation. Viral infectivity was only detectable in the water samples collected immediately after mixing the virus in water containing the water fleas, while no virus was detectable in any of the water fleas or water samples collected afterwards. Conclusions: Water fleas are able to accumulate AIV from surrounding water based upon the qRRT-PCR detection of viral RNA. Additional studies are necessary to investigate the inactivation potential of water fleas on viral infectivity.

In order to investigate the potential role of mussels as a vector of influenza A viruses, we exposed zebra mussels (Dreissena polymorpha) to natural lake water containing a low pathogenic H5N1 avian influenza virus. Mussels were kept in water containing virus for 48 hr, then transferred into fresh water for another 14 days. Virus detection in mussels and water samples was performed by quantitative real-time reverse transcriptase-PCR (qRRT-PCR) and egg culture methods. Virus uptake was detected in all of the mussel groups that were exposed to virus. Even after 14 days in fresh water, virus could still be detected in shellfish material by both qRRT-PCR and egg culture methods. The present study demonstrates that zebra mussels are capable of accumulating influenza A viruses from the surrounding water and that these viruses remain in the mussels over an extended period of time.

A germ carrier technique was adapted for the determination of the persistence of influenza viruses in moist environments. The technique was employed with 3 low pathogenic avian influenza viruses (H4N6, H5N1, and H6N8), one human influenza virus (H1N1), and two model viruses (NDV and ECBO) in lake water at five different temperatures (30, 20, 10, 0, and -10°C). Viral quantitation was carried out at regular intervals on cell culture for a maximum duration of 16 weeks. Serial data were analyzed by linear regression model to calculate T-90 values (time required for one log reduction in the virus titer). Persistence of all of the viruses was highest at -10°C followed by 0, 10, 20, and 30°C. At -10°C, the single freeze-thaw cycle resulted in an abrupt decline in the virus titer, followed by long term persistence. Generally, influenza viruses persisted shorter than model viruses while ECBO has the highest survival time in lake water. Individual influenza viruses differed in their persistence at all temperatures. The findings of the present study suggest that AIV can remain infectious in lake water for extended periods of time at low temperatures.

The incidence of infectious diseases in the United States has been increasing since 1980. Re-emergent conditions, multidrug-resistant bacteria, newly identified infections, and bioterrorism have prompted public health surveillance and control initiatives, including the use of telehealth technology. Infectious diseases, such as West Nile Virus, pose a particular threat to rural areas, where access to infectious disease specialists (IDS) is limited. Initial, in-patient IDS consultations are reimbursed by Centers for Medicare & Medicaid Services for in-person and for telehealth services. Follow-up consultation and subsequent care visits are reimbursed when delivered via in-person care, but not reimbursed when delivered via telehealth. The purpose of this study is to investigate the efficacy of telehealth technology (interactive videoconferencing) in providing timely, efficient, and prudent infectious disease care for rural patients. We conducted a retrospective, comparative review of medical records (n = 107) from inpatients at a metropolitan hospital (n = 59) in a rural state who received in-person IDS treatment, with records from inpatients at nonmetropolitan, rural, and frontier hospitals (n = 48) in the same state who received telehealth IDS treatment. Outcome measures, including number of days hospitalized, number of days receiving intravenous antibiotic, survival, and transfer to another hospital, were compared for three commonly occurring infectious diseases: neutropenic fever, bacterial pneumonia, or bacterial wound infection. Patients treated via telehealth had fewer days on antibiotics and fewer days hospitalized than patients treated via in-person intervention. Survival rates did not differ significantly between groups, but were lower for telehealth patients. Fewer in-person patients required transfer to hospitals offering a higher level of care. Ninety percent of telehealth patients were able to remain at their local hospital for treatment. Results were statistically significant only for selected outcomes and conditions. IDS treatment for the conditions studied is equally effective when delivered via telehealth as when delivered via in-person methods.

We describe a clonal outbreak of quinolone-resistant Haemophilus influenzae (QRHI) from an affiliated long-term care facility (LTCF-A); the outbreak was associated with the clinical use of levofloxacin, which was determined to be a risk factor for acquisition of QRHI. The minimum inhibitory concentration to which 90% of isolates were susceptible (MIC90), as determined by broth microdilution, was >4 microg/mL for levofloxacin, >2 microg/mL for moxifloxacin, >2 microg/mL for gatifloxacin, and 8 microg/mL for gemifloxacin. The MIC90, as determined by Etest (AB Biodisk), was >32 microg/mL for levofloxacin, ciprofloxacin, moxifloxacin, and gatifloxacin. Having been a resident at LTCF-A and having chronic obstructive pulmonary disease were significant risk factors for acquisition of QRHI at our 500-bed hospital (New York Hospital Queens). All QRHI isolates were found to be genetically related by pulsed-field gel electrophoresis, were nontypeable, were susceptible to ceftriaxone and azithromycin, and were negative for beta -lactamase production. Emphasis on patient contact and respiratory isolation and placing colonized or infected patients in cohorts yielded a marked reduction in the prevalence of QRHI at LTCF-A.

Purpose: To study the immunosuppressant and immunopotentiating effects of ketoprofen on antibody producing cells.Methods: Mice were given ketoprofen at doses of 1 mg/kg/day and 5 mg/kg/day for seven days. Similarly polyinosinic–polycytidylic acid (Poly IC) and phosphate buffer saline (PBS) were used as positive and negative control, respectively, for seven days. After seven days, the mice were sacrificed and their spleens removed. Simultaneously, blood was withdrawn from the hearts of the mice and serum was separated from the blood. The spleen cells were analyzed by enzyme-linked immunospot (ELISPOT) while the serum was investigated by enzyme-linked immunosorbant (ELISA) to evaluate the effects of ketoprofen on the ability of individual cell to produce antibodies and antibody- mediated immune responses.Results: Ketoprofen significantly (p < 0.001) reduced the ability of individual cells to produce antibodies. There was a significant difference (p < 0.001) in % of spot forming cells of PBS treated negative control group (0.045 %) as against positive control (0.058 %), 1 mg ketoprofen /kg/day (0.037 %) and 5 mg ketoprofen/kg/day (0.032 %) treated groups. The results of ELISA showed a significant (p < 0.005) difference in the absorbance values between negative control, positive control, ELISA control and ketoprofen treated groups. Absorbance was significantly (p < 0.005) reduced in ketoprofen-treated groups.Conclusion: The ability of an individual cell to produce antibodies and antibody-mediated immune responses is suppressed by ketoprofen, suggesting that it is immunosuppressive, and thus indicating its potential application in patients with auto-immune disorders.Keywords: Ketoprofen, Immunomodulatory, Immunosupressive, Antibody, Spot-forming cells, Polyinosinic–polycytidylic acid

The interaction of nickel nanoparticles with Escherichia coli (E. coli) bacteria has been studied. The nickel nanoparticles have been fabricated by continuous wave laser ablation of nickel target and their properties are studied using different characterization techniques. The antibacterial activity of nickel nanoparticles was checked against E. coli bacteria. Escherichia coli were cultured in nutrients broth and different concentrations of nickel nanoparticles were added to bacterial culture solution to investigate the interaction of nickel nanoparticles with bacteria and to check toxicity of the nickel nanoparticles against E. coli. The fabricated Ni nanoparticles have exhibited considerable antimicrobial activity against E. coli.

Background: Newcastle disease (ND) is one of the most deadly diseases of poultry around the globe. The disease is endemic in Pakistan and recurrent outbreaks are being reported regularly in wild captive, rural and commercial poultry flocks. Though, efforts have been made to characterize the causative agent in some of parts of the country, the genetic nature of strains circulating throughout Pakistan is currently lacking. Material and methods: To ascertain the genetics of NDV, 452 blood samples were collected from 113 flocks, originating from all the provinces of Pakistan, showing high mortality (30 – 80%). The samples represented domesticated poultry (broiler, layer and rural) as well as wild captive birds (pigeons, turkeys, pheasants and peacock). Samples were screened with real-time PCR for both matrix and fusion genes (1792 bp), positive samples were subjected to amplification of full fusion gene and subsequent sequencing and phylogenetic analysis. Results: The deduced amino acid sequence of the fusion protein cleavage site indicated the presence of motif (112 RK/RQRR↓F117) typical for velogenic strains of NDV. Phylogenetic analysis of hypervariable region of the fusion gene indicated that all the isolates belong to lineage 5 of NDV except isolates collected from Khyber Pakhtunkhwa (KPK) province. A higher resolution of the phylogenetic analysis of lineage 5 showed the distribution of Pakistani NDV strains to 5b. However, the isolates from KPK belonged to lineage 4c; the first report of such lineage from this province. Conclusions: Taken together, data indicated the prevalence of multiple lineages of NDV in different poultry population including wild captive birds. Such understanding is crucial to underpin the nature of circulating strains of NDV, their potential for interspecies transmission and disease diagnosis and control strategies. Peer Reviewed publishedVersion